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Keratins and disease ata glance

Rebecca L. Haines and E. BirgitteLane*

Epithelial Biology Group, Institute of Medical Biology,Immunos, Singapore

*Author for correspondence (birgit.lane@imb.a-star.edu.sg)

Journal of Cell Science 125, 3923–3928

� 2012. Published by The Company of Biologists Ltd

doi: 10.1242/jcs.099655

Keratins are cytoskeletal filament-forming

proteins found in skin and other epithelial

(sheet) tissues (Table 1). Keratins (type I and

II), and other highly related (types III–VI)

intermediate filament or nanofilament

proteins, used to be thought of as very inert

because they form filament networks that are

not easily disrupted, in contrast to actin and

microtubule cytoskeleton systems, which

can be rapidly and completely dismantled

by many drugs. The identification of

causative pathogenic mutations in the

keratin genes KRT5 and KRT14 in the

human blistering skin disease epidermolysis

bullosa simplex (EBS) changed this view

and demonstrated the importance of these

proteins for maintaining tissue resilience

(Bonifas et al., 1991; Coulombe et al.,

1991; Lane et al., 1992). As the first-

reported disease caused by keratin

mutations, EBS set the pattern for all the

other keratin disorders, as well as several

non-keratin genetic skin diseases. Many

‘keratinopathies’ have now been identified

with a variety of disease phenotypes,

often predicted by tissue-specific

keratin expression, with pathology usually

involving some form of tissue fragility

(Szeverenyi et al., 2008; see poster).

By reviewing the essential elements of

keratin protein structure and filament

assembly, we consider how keratin

filaments provide tissues with mechanical

resilience, and how mutations might impact

upon these processes and compromise tissue

function. The rare human keratinopathies

have provided important clues to keratin

function, revealing the potential roles for

keratins in many types of stress response,

and even effects on membrane trafficking

and cell and tissue differentiation.

This knowledge has begun to inform

development of therapeutic options for

keratin diseases, and successful first-in-man

siRNA trials have been performed. We

conclude by considering the issues and

challenges now facing the field, and how

they are or can be addressed.

Keratin protein and filamentstructureThe keratin family consists of 54 proteins

(Schweizer et al., 2006) in two families

(types I, acidic, and II, neutral or basic), each

being expressed in a defined sequence during

differentiation. The greatest expression

diversity lies in skin (see poster). Keratin

proteins can be divided into three functional

groups: ‘simple’ keratins, expressed in

embryonic and one-layered epithelia,

including liver, intestine and glandular

secretory cells; ‘barrier’ keratins, expressed

(See poster insert)

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in multilayered stratified squamous epithelia,

such as the epidermis of the skin; and the

harder ‘structural’ keratins that form hair and

nail (Table 1).

Like all intermediate filaments, a keratin

protein has three clear domains: a central

a-helical rod domain, flanked by non-

helical head and tail regions that contain

most phosphorylation sites (Geisler et al.,

1982; Omary et al., 2006). Keratins begin

assembly as heterodimers of one type I and

one type II keratin monomer (Hatzfeld

and Weber, 1990; reviewed by Herrmann

and Aebi, 2000), with rod domains aligned

in parallel and in register. These dimers

form antiparallel tetramers by overlapping

the N-terminal half of their rod domains.

Tetramers then assemble into ‘unit length

filaments’ that are 60 nm in length (see

poster). These rapidly assemble end-to-

end, within 10 seconds in vitro (Herrmann

et al., 2002), forming rope-like

nanofilaments that are ,10 nm thick. The

antiparallel nature of the tetramers results

in filaments that do not have polarity,

which is again in contrast to actin filaments

or microtubules, and implies that keratin

filaments cannot function as unidirectional

tracks for molecular motors. At either end

of the rod domain are the highly conserved

helix initiation and termination motifs,

which are hotspots for the most severe

disease mutations (Lane and McLean,

2004; Uitto et al., 2007).

Dynamic behaviour of keratinsThe cytoplasmic ropes of keratin form

branching bundles in the cell and anchor

into junctions around the cell perimeter.

Keratins link through desmoplakin into

desmosomes (Kouklis et al., 1994), which

connect neighbouring cells, and through

plectin (Andra et al., 2003) into

hemidesmosomes, which connect cells to

their attachment substrates. This filament–

junction network anchors cells in three

dimensions through the epithelium.

However, the keratin filament cytoskeleton

must be dynamic, plastic and flexible to

allow cells to proliferate during growth and

to migrate during wound healing. Dynamic

assembly and disassembly of filaments is

also needed to allow epithelial cells to

maintain an intact network while they alter

their keratin expression profile during

differentiation or in response to stress. For

example, in the epidermis, K5 and K14 are

the major keratins synthesised in basal cells

(which can proliferate), whereas K1 and

K10 are expressed in suprabasal cells (which

are ‘locked’ into differentiation). However,

keratin proteins have a long half-life, around

4 days for simple keratins (Denk et al.,

1987), and K5 and K14 can persist in

suprabasal cells (see poster) alongside K1

and K10 (Kartasova et al., 1993; Eriksson

et al., 2009; Windoffer et al., 2011). A

gradual transition, rather than complete

disassembly and re-assembly of the

network, preserves the structural integrity of

the filament network during differentiation.

Where do keratins assemble in the cell?

Intermediate filaments do not have defined

organizing centres as microtubules do;

they can be synthesised throughout the

cell as tiny subfilamentous particles that

occur throughout the cytoplasm (Liovic

et al., 2003). The unit length filament (see

poster) might be the nucleation precursor,

but this has not been confirmed as its size

(60 nm) is below the resolution limit

of standard light microscopy. New

super-resolution microscopy techniques

(reviewed by Schermelleh et al., 2010)

could soon shed light on this. Turnover

might be accelerated at the cell periphery,

and a model of filament assembly has been

Table 1. Expression of keratin proteins in epithelial tissues

Keratin Epithelial tissue Polymerisation partner in vivo

Type ISimple K18 Simple epithelia (e.g. liver, pancreas, colon, lung) K8, K7

K20 Simple epithelia, especially gastrointestinal K8, (K7)Barrier K9 Stratified cornifying epithelia; palm, sole (K1)

K10 Stratified cornifying epithelia, suprabasal K1K12 Stratified epithelia; cornea K3K13 Stratified epithelia, non-cornifying; suprabasal K4K14 Stratified and complex epithelia; basal K5K15 Stratified epithelia (K5)K16 Stratified epithelia; induced during stress, fast turnover;

suprabasalK6a

K17 Stratified epithelia; induced during stress, fast turnover K6bK19 Simple and stratified epithelia K8K23, K24 Epithelia

Structural K25, K26, K27, K28 Stratified epithelia: hair follicle sheathK31, K32, K33a, K33b, K34, K35,

K36, K37, K38, K39, K40Stratified epithelia: hair, hard structures

Type IISimple K7, K8 Simple epithelia K18Barrier K1 Stratified epithelia, cornifying, suprabasal K10

K2 Stratified cornifying epithelia, late suprabasal (K10)K3 Stratified epithelia, cornea K12K4 Stratified epithelia, non-cornifying; suprabasal K13K5 Stratified and complex epithelia; basal cells K14, (K15)K6a Stratified epithelia; induced during stress, fast turnover K16K6b Stratified epithelia; induced during stress, fast turnover K17K6c EpitheliaK76 Stratified cornifying epithelia, oral, suprabasal (K10)K78, K79, K80 Epithelia

Structural K75 Stratified epithelia: hair follicleK71, K72, K73, K74 Stratified epithelia: hair follicle sheathK81, K82, K83, K84, K85, K86 Stratified epithelia: hair, hard structures

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recently described that proposes that

filaments are nucleated in the cell

periphery at focal adhesions, before

elongating and then integrating into the

pre-existing peripheral network (Windoffer

et al., 2011).

A probable cofactor for localising the

assembling keratins is the linker protein

plectin because plectin isoforms bind actin

or intermediate filaments at different cell

locations. Plectin is known to influence

the formation and dynamics of vimentin

filaments (the intermediate filament

protein in fibroblasts) (Spurny et al.,

2008; Kostan et al., 2009), and plectin

has also been implicated in the actin-

dependent inward movement of keratins in

the cell periphery (Kolsch et al., 2009).

Keratin assembly at focal adhesions

(Windoffer et al., 2006) might also

depend on phosphorylation by the p38

mitogen-activated protein kinases, as

inhibition of these kinases prevents the

formation of filament precursors in the cell

periphery (Woll et al., 2007).

A wide range of post-translational

modifications have been described

on keratins, including phosphorylation

to ubiquitylation, sumoylation and

acetylation (Omary et al., 2006; Ku et al.,

2010; Srikanth et al., 2010; Snider et al.,

2011), which are likely to modulate the

solubility of keratins in specific situations.

The most well-documented modification is

phosphorylation, with many sites known

on simple keratins (reviewed by Omary

et al., 2006) but fewer defined on barrier

keratins. Phosphorylation of keratin

filaments by protein kinase Cf is required

for K8 and K18 filament remodelling in

response to shear stress (Flitney et al.,

2009; Sivaramakrishnan et al., 2009).

Considering all the above, even a single

amino acid change in keratins can interrupt

cell function in many ways, by altering

their post-translational modifications, their

integration into junctions or their filament

assembly kinetics (Herrmann et al., 2002;

Owens et al., 2004) to yield a less-stable

filament network and cause disease. For

example, some of the severe K14

mutations that cause Dowling–Meara

EBS, such as the hotspot mutation

p.Arg125Pro, result in the formation of

cytoplasmic keratin aggregates (Ishida-

Yamamoto et al., 1991). In vitro, this

mutation reduces the ability of

reconstituted filaments to bundle under

cross-linking conditions (Ma et al., 2001).

The impact of mutations on the mechanical

resilience of epithelial tissues is discussedbelow.

Keratins provide epithelia withmechanical resilienceThe keratin filament network can withstand

significant mechanical forces, particularly instratified epithelia such as the epidermis(Beriault et al., 2012). By conferring

mechanical continuity across an epithelialsheet, the keratin–desmosome and keratin–hemidesmosome network generates

mechanical resilience across the tissue, asthe network will dissipate mechanical stressaway from a source into the surroundingepithelium. Investigations of single-filament

mechanical properties show that keratinfilaments are flexible and tough, being lessrigid than actin filaments at low strains, and

less brittle than microtubules at high strains,where they show strain hardening (Janmeyet al., 1991; Kreplak and Fudge, 2007).

Bundling of keratin filaments appears toincrease when cells are subjected to shearstress (Flitney et al., 2009).

The tissue fragility observed in many

keratin diseases reflects the crucial role ofkeratins in providing mechanical stability tocells and tissues (McLean and Moore, 2011).

However, studies suggest that mutations inkeratins do not reduce mechanical resiliencesimply by preventing filament formation. A

keratinocyte cell line expressing a GFP-tagged K14 carrying the p.Arg125Promutation can withstand uni-directional

stretch (greater than 100%) withoutsignificant damage or cell death as much ascells expressing wild-type filaments can(Fudge et al., 2008; Beriault et al., 2012).

However, the cells expressing mutantkeratins are much less able to survive arepetitive stretch than those expressing wild-

type keratins (Russell et al., 2004). It appearslikely that mutations alter the dynamics offilament formation, giving rise to a keratin

network that is less stable. Furthermore, celllines generated from EBS patients withmutations in K5 or K14 migrate faster intissue culture scratch wound assays than their

wild-type counterparts, perhaps because thenetwork is more dynamic, allowing the cellto re-organise its keratin filaments more

quickly for migration (Morley et al., 2003).The presence of keratin mutations, or areduction in keratin expression, also reduces

expression of desmosome components andcytoskeletal linker proteins (Long et al.,2006; Liovic et al., 2009; Wagner et al.,

2012). This suggests an alternative diseasemechanism in which keratin mutations mightresult in tissue fragility due to a reduction in

junction proteins, leading to decreasedstability of the tissue.

Keratin disease phenotypes suggestthat there are multiple functionsfor keratinsEvidence from human keratin diseases thatare not associated with tissue fragility, as

well as data from experimental models suchas knockout and transgenic mice, hassuggested that keratins are essential formany different cellular processes, including

non-mechanical stress responses, organellepositioning and tissue differentiation (Guand Coulombe, 2007; Omary et al., 2009;

Toivola et al., 2010). These are discussed inthe following sections.

Mutant keratins alter stress response

Expression of many keratins is upregulated

in response to stress, suggesting animportant role for these proteins in thestress response (reviewed by Toivola et al.,2010). In particular, the expression of

keratins K6a, K6b, K16 and K17 isinduced by inflammatory cytokines, or inresponse to wound healing, and oxidative

or UV stress (Freedberg et al., 2001;DePianto and Coulombe, 2004).

Mutations in the simple epithelialkeratins K8 and K18 have been identified

as risk factors for some patients withinflammatory bowel disease (Owens et al.,2004) or liver disease (Ku et al., 2003).

Loss of mechanical resilience in theintestine (required to withstand peristalticmovement during digestion) caused bykeratin mutations might initiate tissue

damage (Owens and Lane, 2004). Bycontrast, liver is less mechanicallystressed but is vulnerable to damage by

toxins such as alcohol, and, in liver, keratinmutations reduce the tolerance to suchassaults (Ku et al., 2007; Omary et al.,

2009). This might help explain whymutations in these keratins are silent inmost individuals, but still predisposecarriers to liver injury mediated by toxins

or viruses, which constitute situations ofcell stress (Omary et al., 2009). It has beensuggested K8 acts as a ‘phosphate sponge’,

undergoing hyperphosphorylation, whichabsorbs phosphorylated stress-activatedprotein kinase (SAPK) activity, thus

preventing apoptosis (see poster; Ku andOmary, 2006).

Mutations in K5 and K14 that causeEBS also alter the response of a cell to

stress. Cells expressing mutated K5 or K14show amplified and accelerated SAPKsignalling in response to external stresses

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(D’Alessandro et al., 2002), and the

constitutive upregulation of extracellular-signal-regulated kinase (ERK) signalling inthese cells contributes to their increased

resistance to apoptosis (Russell et al.,2010). The targeting of keratin filamentsfor degradation is increased during stressand by the presence of mutated or

misfolded keratins that cannot integrateefficiently into the keratin network(Jaitovich et al., 2008; Loffek et al.,

2010; Na et al., 2010; Rogel et al., 2010).

Keratins and organelle transport

Mutations in some keratins have revealed a

role for these proteins in certain membranetrafficking events (see poster) (Kumemuraet al., 2004; Toivola et al., 2005; Kumemura

et al., 2008). A group of rare keratindiseases, including Dowling–Degosdisease, that are caused by mutations in K5or K14 have a skin pigment phenotype

(patches of hyper- and hypo-pigmentedskin; see poster) that is not directly linkedto skin blistering (when present) (Uttam

et al., 1996; Betz et al., 2006; Lugassy et al.,2006). Melanosomes (pigment-containinggranules) are produced by melanocytes and

transferred into basal keratinocytes wherethey are arranged in a distal cap over thenucleus. A defect in the transfer of

melanosomes, or their arrangement inkeratinocytes, can lead to changes in skinpigmentation. Although the mechanism bywhich mutations in epidermal keratins alter

melanosome arrangement is not fullyunderstood, it might involve an interactionof K5 with the chaperone HSC70, which is

involved in vesicle uncoating (Planko et al.,2007).

Tissue differentiation is affected bykeratin expression

The tissue-specific expression pattern ofkeratins is set early in development and

differentiation and is therefore tightlycontrolled. Histopathologists havehistorically used keratin expression inepithelial tumours to identify the tissue of

origin (Lane and Alexander, 1990), andmore recently as prognostic markers(reviewed by Karantza, 2011). Changing

the keratins that are expressed in a cell canhave wide-ranging effects; these might berelated to a change in cell fate that is

reflected in the altered keratin profile andmight lead to a subsequent alteration inphysical properties (discussed in Owens and

Lane, 2003). For example, mice expressingK1 in pancreatic b cells develop diabetescharacterised by a reduction of insulin-

secretory vesicles (Blessing et al., 1993).Deletion of K17 in a mouse model leads to

defects in hair follicle cycling, wound repair(through a decrease in cell size and proteinsynthesis), and an altered inflammatorycytokine profile (Kim et al., 2006; Tong

and Coulombe, 2006; DePianto et al., 2010).Although interactions of K17 with theadaptor protein 14-3-3s and TRADD

(tumour necrosis factor receptor type 1-associated death domain) suggest potentialmechanisms by which loss of K17 could

give rise to some of these phenotypes, itseems likely that deletion of K17 has a moregeneral effect on tissue differentiation.Studies of ‘unnatural’ keratin pairs, such as

K5 paired with K16 or K18 rather than itsnormal partner, K14, suggest that theassembly and mechanical properties of the

filaments can be drastically altered bychanging the type I or type II keratinpresent (Yamada et al., 2002; Lee and

Coulombe, 2009). This is reflected inthe mouse knockout of K14, in whichexpression of K16 or K18 only leads to

partial rescue of the phenotype, to differingdegrees (Hutton et al., 1998; Paladini andCoulombe, 1999), indicating that expressionof the correct keratins is crucial for normal

tissue function. The correct expressionof keratins is also important in thedevelopment and maintenance of

epidermal appendages (hair follicles andsebaceous glands). Mutations in the keratinsexpressed in these appendages lead to

various defects including the hair fragilitysyndrome monilethrix, or pachyonychiacongenita, a disorder characterised bynail dystrophy, painful palmoplantar

keratoderma, and either hair-follicle-associated cysts or cell fragility in mucoussurfaces (McLean et al., 2005; Schweizer

et al., 2007) (see poster).

Understanding the precise role of keratinsin these processes, and how mutations give

rise to the defects described will be crucialto the development of therapies for thekeratin diseases.

Opportunities for therapySo far, therapeutic approaches to keratindiseases have focussed on two different

areas, (i) genetic ablation of the mutantprotein, and (ii) small-molecule therapies tostabilise the keratin network in some way. A

successful trial of small interfering RNA(siRNA) against mutant K6a in apachyonychia congenita patient recently

demonstrated the feasibility of the firsttherapeutic route (Leachman et al., 2010),and without doubt further siRNA-based

approaches will follow (Atkinson et al.,2011; Liao et al., 2011). However,

significant concerns have been expressedregarding potential off-target effects ofsiRNA (Jackson and Linsley, 2010). Aschanges in keratin expression can have

dramatic effects on tissue differentiation(see above), attempts to alter keratinexpression for therapeutic purposes must

be approached with caution. Nevertheless,small-molecule therapies are also beinginvestigated to manipulate keratin levels in

the cell. Several small molecules have nowbeen shown to modulate keratin expression,and they either alter the expression ofseveral keratins or target a specific keratin.

They include statins, which moderatelydownregulate the activity of the promoterfor K6a (Kerns et al., 2007; Torma, 2011;

Zhao et al., 2011). In severe keratindisorders, where aggregates are observed,it might be possible to ameliorate the

symptoms by reducing the amount ofaggregation with the use of chemicalchaperones, which could ‘clear the way’

for the keratin filament cytoskeleton toreform correctly (Loffek et al., 2010;Chamcheu et al., 2011).

PerspectivesKeratin filaments have a crucial functionin providing mechanical resilience to

epithelial tissues. Associated with thisfunction, keratins are involved in the cellstress response, tissue differentiation and

organelle transport. Despite significanteffort, the details of the substructure ofkeratin filaments, and in particular the roleof the head and tail domains, are poorly

understood (Strelkov et al., 2003), as is thecontrol of filament assembly and dynamicswithin the cell. One model of filament

dynamics, in which filaments are nucleatedin the cell periphery and elongate andmature as they move towards the centre of

the cell, was presented above. However,filament dynamics are likely to be differentin a migrating or dividing cell, where thekeratin cytoskeleton is highly dynamic,

compared with cells in an intact tissue,where filaments appear to be stable and areanchored at cell junctions. A combination

of localised highly dynamic filament re-organisation [as has been observed forvimentin (Helfand et al., 2011)] and some

form of subunit exchange within thenetwork seems more likely, but themechanisms by which keratin dynamics

are controlled are not yet understood.Disease-causing keratin mutations mightprovide essential information in this area,

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particularly in explaining how changing

filament formation and dynamics give rise

to fragile epithelial tissues. In order to

address these issues, it will be necessary to

improve laboratory models of keratin

diseases to better reflect the physiological

condition. Understanding the connection

between the role of keratin filaments in

providing mechanical resilience and the

apparently unrelated phenotypes that are

observed in certain keratin diseases is a

priority, and further investigations in this

area will hopefully yield new therapeutic

interventions for keratin diseases.

Acknowledgements

The authors would like to thank John

Common for critical reading of the

manuscript, and John Common, Declan

Lunny and Graham Wright (IMB

Microscopy Unit) for assistance in

producing images.

Funding

Work in the authors’ laboratory is supported

by the Biomedical Research Council,

Agency for Science, Technology and

Research (A*STAR), Singapore.

A high-resolution version of the poster is available for

downloading in the online version of this article at

jcs.biologists.org. Individual poster panels are available

as JPEG files at http://jcs.biologists.org/lookup/suppl/

doi:10.1242/jcs.099655/-/DC1

ReferencesAndra, K., Kornacker, I., Jorgl, A., Zorer, M.,

Spazierer, D., Fuchs, P., Fischer, I. and Wiche, G.

(2003). Plectin-isoform-specific rescue of

hemidesmosomal defects in plectin (-/-) keratinocytes. J.

Invest. Dermatol. 120, 189-197.

Atkinson, S. D., McGilligan, V. E., Liao, H.,

Szeverenyi, I., Smith, F. J. D., Moore, C. B. T. and

McLean, W. H. I. (2011). Development of allele-specific

therapeutic siRNA for keratin 5 mutations in

epidermolysis bullosa simplex. J. Invest. Dermatol. 131,

2079-2086.

Beriault, D. R., Haddad, O., McCuaig, J. V., Robinson,

Z. J., Russell, D., Lane, E. B. and Fudge, D. S. (2012).

The mechanical behavior of mutant K14-R125P keratin

bundles and networks in NEB-1 keratinocytes. PLoS ONE

7, e31320.

Betz, R. C., Planko, L., Eigelshoven, S., Hanneken, S.,

Pasternack, S. M., Bussow, H., Van Den Bogaert, K.,

Wenzel, J., Braun-Falco, M., Rutten, A. et al. (2006).

Loss-of-function mutations in the keratin 5 gene lead to

Dowling-Degos disease. Am. J. Hum. Genet. 78, 510-519.

Blessing, M., Ruther, U. and Franke, W. W. (1993).

Ectopic synthesis of epidermal cytokeratins in pancreatic

islet cells of transgenic mice interferes with cytoskeletal

order and insulin production. J. Cell Biol. 120, 743-755.

Bonifas, J. M., Rothman, A. L. and Epstein, E. H., Jr

(1991). Epidermolysis bullosa simplex: evidence in two

families for keratin gene abnormalities. Science 254,

1202-1205.

Chamcheu, J. C., Navsaria, H., Pihl-Lundin, I., Liovic,

M., Vahlquist, A. and Torma, H. (2011). Chemical

chaperones protect epidermolysis bullosa simplex

keratinocytes from heat stress-induced keratin

aggregation: involvement of heat shock proteins and

MAP kinases. J. Invest. Dermatol. 131, 1684-1691.

Coulombe, P. A., Hutton, M. E., Letai, A., Hebert, A.,

Paller, A. S. and Fuchs, E. (1991). Point mutations in

human keratin 14 genes of epidermolysis bullosa simplex

patients: genetic and functional analyses. Cell 66, 1301-

1311.

D’Alessandro, M., Russell, D., Morley, S. M., Davies,

A. M. and Lane, E. B. (2002). Keratin mutations of

epidermolysis bullosa simplex alter the kinetics of stress

response to osmotic shock. J. Cell Sci. 115, 4341-4351.

Denk, H., Lackinger, E., Zatloukal, K. and Franke,

W. W. (1987). Turnover of cytokeratin polypeptides in

mouse hepatocytes. Exp. Cell Res. 173, 137-143.

DePianto, D. and Coulombe, P. A. (2004). Intermediate

filaments and tissue repair. Exp. Cell Res. 301, 68-76.

DePianto, D., Kerns, M. L., Dlugosz, A. A. and

Coulombe, P. A. (2010). Keratin 17 promotes epithelial

proliferation and tumor growth by polarizing the immuneresponse in skin. Nat. Genet. 42, 910-914.

Eriksson, J. E., Dechat, T., Grin, B., Helfand, B.,

Mendez, M., Pallari, H.-M. and Goldman, R. D. (2009).

Introducing intermediate filaments: from discovery to

disease. J. Clin. Invest. 119, 1763-1771.

Flitney, E. W., Kuczmarski, E. R., Adam, S. A. and

Goldman, R. D. (2009). Insights into the mechanical

properties of epithelial cells: the effects of shear stress on

the assembly and remodeling of keratin intermediate

filaments. FASEB J. 23, 2110-2119.

Freedberg, I. M., Tomic-Canic, M., Komine, M. and

Blumenberg, M. (2001). Keratins and the keratinocyteactivation cycle. J. Invest. Dermatol. 116, 633-640.

Fudge, D., Russell, D., Beriault, D., Moore, W., Lane,

E. B. and Vogl, A. W. (2008). The intermediate filament

network in cultured human keratinocytes is remarkably

extensible and resilient. PLoS ONE 3, e2327.

Geisler, N., Kaufmann, E. and Weber, K. (1982).

Proteinchemical characterization of three structurally

distinct domains along the protofilament unit of desmin

10 nm filaments. Cell 30, 277-286.

Gu, L.-H. and Coulombe, P. A. (2007). Keratin function

in skin epithelia: a broadening palette with surprising

shades. Curr. Opin. Cell Biol. 19, 13-23.

Hatzfeld, M. and Weber, K. (1990). The coiled coil of in

vitro assembled keratin filaments is a heterodimer of type

I and II keratins: use of site-specific mutagenesis and

recombinant protein expression. J. Cell Biol. 110, 1199-

1210.

Helfand, B. T., Mendez, M. G., Murthy, S. N. P.,

Shumaker, D. K., Grin, B., Mahammad, S., Aebi, U.,

Wedig, T., Wu, Y. I., Hahn, K. M. et al. (2011).

Vimentin organization modulates the formation of

lamellipodia. Mol. Biol. Cell 22, 1274-1289.

Herrmann, H. and Aebi, U. (2000). Intermediate

filaments and their associates: multi-talented structuralelements specifying cytoarchitecture and cytodynamics.

Curr. Opin. Cell Biol. 12, 79-90.

Herrmann, H., Wedig, T., Porter, R. M., Lane, E. B.

and Aebi, U. (2002). Characterization of early assembly

intermediates of recombinant human keratins. J. Struct.

Biol. 137, 82-96.

Hutton, E., Paladini, R. D., Yu, Q. C., Yen, M.,

Coulombe, P. A. and Fuchs, E. (1998). Functional

differences between keratins of stratified and simple

epithelia. J. Cell Biol. 143, 487-499.

Ishida-Yamamoto, A., McGrath, J. A., Chapman, S. J.,

Leigh, I. M., Lane, E. B. and Eady, R. A. (1991).

Epidermolysis bullosa simplex (Dowling-Meara type) is a

genetic disease characterized by an abnormal keratin-

filament network involving keratins K5 and K14. J. Invest.

Dermatol. 97, 959-968.

Jackson, A. L. and Linsley, P. S. (2010). Recognizing

and avoiding siRNA off-target effects for target

identification and therapeutic application. Nat. Rev.

Drug Discov. 9, 57-67.

Jaitovich, A., Mehta, S., Na, N., Ciechanover, A.,

Goldman, R. D. and Ridge, K. M. (2008). Ubiquitin-

proteasome-mediated degradation of keratin intermediate

filaments in mechanically stimulated A549 cells. J. Biol.

Chem. 283, 25348-25355.

Janmey, P. A., Euteneuer, U., Traub, P. and Schliwa,

M. (1991). Viscoelastic properties of vimentin compared

with other filamentous biopolymer networks. J. Cell Biol.

113, 155-160.

Karantza, V. (2011). Keratins in health and cancer: morethan mere epithelial cell markers. Oncogene 30, 127-138.

Kartasova, T., Roop, D. R., Holbrook, K. A. and

Yuspa, S. H. (1993). Mouse differentiation-specifickeratins 1 and 10 require a preexisting keratin scaffoldto form a filament network. J. Cell Biol. 120, 1251-1261.

Kerns, M. L., DePianto, D., Dinkova-Kostova, A. T.,

Talalay, P. and Coulombe, P. A. (2007).Reprogramming of keratin biosynthesis by sulforaphanerestores skin integrity in epidermolysis bullosa simplex.Proc. Natl. Acad. Sci. USA 104, 14460-14465.

Kim, S., Wong, P. and Coulombe, P. A. (2006). Akeratin cytoskeletal protein regulates protein synthesis andepithelial cell growth. Nature 441, 362-365.

Kolsch, A., Windoffer, R. and Leube, R. E. (2009).Actin-dependent dynamics of keratin filament precursors.Cell Motil. Cytoskeleton 66, 976-985.

Kostan, J., Gregor, M., Walko, G. and Wiche, G.

(2009). Plectin isoform-dependent regulation of keratin-integrin alpha6beta4 anchorage via Ca2+/calmodulin. J.

Biol. Chem. 284, 18525-18536.

Kouklis, P. D., Hutton, E. and Fuchs, E. (1994). Makinga connection: direct binding between keratin intermediatefilaments and desmosomal proteins. J. Cell Biol. 127,1049-1060.

Kreplak, L. and Fudge, D. (2007). Biomechanicalproperties of intermediate filaments: from tissues tosingle filaments and back. Bioessays 29, 26-35.

Ku, N.-O. and Omary, M. B. (2006). A disease- andphosphorylation-related nonmechanical function forkeratin 8. J. Cell Biol. 174, 115-125.

Ku, N. O., Darling, J. M., Krams, S. M., Esquivel,C. O., Keeffe, E. B., Sibley, R. K., Lee, Y. M., Wright,

T. L. and Omary, M. B. (2003). Keratin 8 and 18mutations are risk factors for developing liver disease ofmultiple etiologies. Proc. Natl. Acad. Sci. USA 100, 6063-6068.

Ku, N.-O., Strnad, P., Zhong, B.-H., Tao, G.-Z. and

Omary, M. B. (2007). Keratins let liver live: Mutationspredispose to liver disease and crosslinking generatesMallory-Denk bodies. Hepatology 46, 1639-1649.

Ku, N.-O., Toivola, D. M., Strnad, P. and Omary, M. B.

(2010). Cytoskeletal keratin glycosylation protectsepithelial tissue from injury. Nat. Cell Biol. 12, 876-885.

Kumemura, H., Harada, M., Omary, M. B., Sakisaka,

S., Suganuma, T., Namba, M. and Sata, M. (2004).Aggregation and loss of cytokeratin filament networksinhibit golgi organization in liver-derived epithelial celllines. Cell Motil. Cytoskeleton 57, 37-52.

Kumemura, H., Harada, M., Yanagimoto, C., Koga,H., Kawaguchi, T., Hanada, S., Taniguchi, E., Ueno, T.

and Sata, M. (2008). Mutation in keratin 18 inducesmitochondrial fragmentation in liver-derived epithelialcells. Biochem. Biophys. Res. Commun. 367, 33-40.

Lane, E. B. and Alexander, C. M. (1990). Use of keratinantibodies in tumor diagnosis. Semin. Cancer Biol. 1, 165-179.

Lane, E. B. and McLean, W. H. I. (2004). Keratins andskin disorders. J. Pathol. 204, 355-366.

Lane, E. B., Rugg, E. L., Navsaria, H., Leigh, I. M.,

Heagerty, A. H. M., Ishida-Yamamoto, A. and Eady,R. A. J. (1992). A mutation in the conserved helixtermination peptide of keratin 5 in hereditary skinblistering. Nature 356, 244-246.

Leachman, S. A., Hickerson, R. P., Schwartz, M. E.,

Bullough, E. E., Hutcherson, S. L., Boucher, K. M.,Hansen, C. D., Eliason, M. J., Srivatsa, G. S.,

Kornbrust, D. J. et al. (2010). First-in-human mutation-targeted siRNA phase Ib trial of an inherited skin disorder.Mol. Ther. 18, 442-446.

Lee, C.-H. and Coulombe, P. A. (2009). Self-organization of keratin intermediate filaments into cross-linked networks. J. Cell Biol. 186, 409-421.

Liao, H., Irvine, A. D., Macewen, C. J., Weed, K. H.,

Porter, L., Corden, L. D., Gibson, A. B., Moore, J. E.,Smith, F. J. D., McLean, W. H. I. et al. (2011).Development of allele-specific therapeutic siRNA inMeesmann epithelial corneal dystrophy. PLoS ONE 6,e28582.

Liovic, M., Mogensen, M. M., Prescott, A. R. and Lane,

E. B. (2003). Observation of keratin particles showing fastbidirectional movement colocalized with microtubules. J.

Cell Sci. 116, 1417-1427.

Journal of Cell Science 125 (17) 3927

Journ

alof

Cell

Scie

nce

Liovic, M., D’Alessandro, M., Tomic-Canic, M.,

Bolshakov, V. N., Coats, S. E. and Lane, E. B. (2009).Severe keratin 5 and 14 mutations induce down-regulationof junction proteins in keratinocytes. Exp. Cell Res. 315,2995-3003.

Loffek, S., Woll, S., Hohfeld, J., Leube, R. E., Has, C.,

Bruckner-Tuderman, L. and Magin, T. M. (2010). Theubiquitin ligase CHIP/STUB1 targets mutant keratins fordegradation. Hum. Mutat. 31, 466-476.

Long, H. A., Boczonadi, V., McInroy, L., Goldberg, M.

and Maatta, A. (2006). Periplakin-dependent re-organisation of keratin cytoskeleton and loss ofcollective migration in keratin-8-downregulatedepithelial sheets. J. Cell Sci. 119, 5147-5159.

Lugassy, J., Itin, P., Ishida-Yamamoto, A., Holland, K.,

Huson, S., Geiger, D., Hennies, H. C., Indelman, M.,

Bercovich, D., Uitto, J. et al. (2006). Naegeli-Franceschetti-Jadassohn syndrome and dermatopathiapigmentosa reticularis: two allelic ectodermal dysplasiascaused by dominant mutations in KRT14. Am. J. Hum.

Genet. 79, 724-730.

Ma, L., Yamada, S., Wirtz, D. and Coulombe, P. A.

(2001). A ‘hot-spot’ mutation alters the mechanicalproperties of keratin filament networks. Nat. Cell Biol.

3, 503-506.

McLean, W. H. I. and Moore, C. B. T. (2011). Keratindisorders: from gene to therapy. Hum. Mol. Genet. 20,R189-R197.

McLean, W. H. I., Smith, F. J. D. and Cassidy, A. J.

(2005). Insights into genotype-phenotype correlation in

pachyonychia congenita from the human intermediatefilament mutation database. J. Investig. Dermatol. Symp.

Proc. 10, 31-36.

Morley, S. M., D’Alessandro, M., Sexton, C., Rugg,

E. L., Navsaria, H., Shemanko, C. S., Huber, M., Hohl,

D., Heagerty, A. I., Leigh, I. M. et al. (2003). Generationand characterization of epidermolysis bullosa simplex celllines: scratch assays show faster migration with disruptivekeratin mutations. Br. J. Dermatol. 149, 46-58.

Na, N., Chandel, N. S., Litvan, J. and Ridge, K. M.

(2010). Mitochondrial reactive oxygen species are

required for hypoxia-induced degradation of keratinintermediate filaments. FASEB J. 24, 799-809.

Omary, M. B., Ku, N.-O., Tao, G.-Z., Toivola, D. M.

and Liao, J. (2006). ‘‘Heads and tails’’ of intermediatefilament phosphorylation: multiple sites and functional

insights. Trends Biochem. Sci. 31, 383-394.

Omary, M. B., Ku, N.-O., Strnad, P. and Hanada, S.

(2009). Toward unraveling the complexity of simpleepithelial keratins in human disease. J. Clin. Invest. 119,1794-1805.

Owens, D. W. and Lane, E. B. (2003). The quest for thefunction of simple epithelial keratins. Bioessays 25, 748-758.

Owens, D. W. and Lane, E. B. (2004). Keratin mutationsand intestinal pathology. J. Pathol. 204, 377-385.Owens, D. W., Wilson, N. J., Hill, A. J. M., Rugg, E. L.,Porter, R. M., Hutcheson, A. M., Quinlan, R. A., van

Heel, D., Parkes, M., Jewell, D. P. et al. (2004). Humankeratin 8 mutations that disturb filament assemblyobserved in inflammatory bowel disease patients. J. Cell

Sci. 117, 1989-1999.Paladini, R. D. and Coulombe, P. A. (1999). Thefunctional diversity of epidermal keratins revealed by thepartial rescue of the keratin 14 null phenotype by keratin16. J. Cell Biol. 146, 1185-1201.Planko, L., Bohse, K., Hohfeld, J., Betz, R. C.,Hanneken, S., Eigelshoven, S., Kruse, R., Nothen,

M. M. and Magin, T. M. (2007). Identification of akeratin-associated protein with a putative role in vesicletransport. Eur. J. Cell Biol. 86, 827-839.Rogel, M. R., Jaitovich, A. and Ridge, K. M. (2010).The role of the ubiquitin proteasome pathway in keratinintermediate filament protein degradation. Proc. Am.

Thorac. Soc. 7, 71-76.Russell, D., Andrews, P. D., James, J. and Lane, E. B.

(2004). Mechanical stress induces profound remodellingof keratin filaments and cell junctions in epidermolysisbullosa simplex keratinocytes. J. Cell Sci. 117, 5233-5243.Russell, D., Ross, H. and Lane, E. B. (2010). ERKinvolvement in resistance to apoptosis in keratinocyteswith mutant keratin. J. Invest. Dermatol. 130, 671-681.Schermelleh, L., Heintzmann, R. and Leonhardt, H.

(2010). A guide to super-resolution fluorescencemicroscopy. J. Cell Biol. 190, 165-175.Schweizer, J., Bowden, P. E., Coulombe, P. A.,Langbein, L., Lane, E. B., Magin, T. M., Maltais, L.,

Omary, M. B., Parry, D. A. D., Rogers, M. A. et al.

(2006). New consensus nomenclature for mammaliankeratins. J. Cell Biol. 174, 169-174.Schweizer, J., Langbein, L., Rogers, M. A. and Winter,H. (2007). Hair follicle-specific keratins and theirdiseases. Exp. Cell Res. 313, 2010-2020.Sivaramakrishnan, S., Schneider, J. L., Sitikov, A.,Goldman, R. D. and Ridge, K. M. (2009). Shear stressinduced reorganization of the keratin intermediatefilament network requires phosphorylation by proteinkinase C zeta. Mol. Biol. Cell 20, 2755-2765.Snider, N. T., Weerasinghe, S. V. W., Iniguez-Lluhı,

J. A., Herrmann, H. and Omary, M. B. (2011). Keratinhypersumoylation alters filament dynamics and is amarker for human liver disease and keratin mutation. J.

Biol. Chem. 286, 2273-2284.Spurny, R., Gregor, M., Castanon, M. J. and Wiche, G.

(2008). Plectin deficiency affects precursor formation anddynamics of vimentin networks. Exp. Cell Res. 314, 3570-3580.Srikanth, B., Vaidya, M. M. and Kalraiya, R. D. (2010).O-GlcNAcylation determines the solubility, filament

organization, and stability of keratins 8 and 18. J. Biol.

Chem. 285, 34062-34071.Strelkov, S. V., Herrmann, H. and Aebi, U. (2003).Molecular architecture of intermediate filaments.Bioessays 25, 243-251.Szeverenyi, I., Cassidy, A. J., Chung, C. W., Lee, B. T. K.,Common, J. E. A., Ogg, S. C., Chen, H., Sim, S. Y., Goh,

W. L. P., Ng, K. W. et al. (2008). The Human IntermediateFilament Database: comprehensive information on a genefamily involved in many human diseases. Hum. Mutat. 29,351-360.Toivola, D. M., Tao, G.-Z., Habtezion, A., Liao, J. and

Omary, M. B. (2005). Cellular integrity plus: organelle-related and protein-targeting functions of intermediatefilaments. Trends Cell Biol. 15, 608-617.Toivola, D. M., Strnad, P., Habtezion, A. and Omary,

M. B. (2010). Intermediate filaments take the heat asstress proteins. Trends Cell Biol. 20, 79-91.Tong, X. and Coulombe, P. A. (2006). Keratin 17modulates hair follicle cycling in a TNFalpha-dependentfashion. Genes Dev. 20, 1353-1364.Torma, H. (2011). Regulation of keratin expression byretinoids. Dermatoendocrinol. 3, 136-140.Uitto, J., Richard, G. and McGrath, J. A. (2007).Diseases of epidermal keratins and their linker proteins.Exp. Cell Res. 313, 1995-2009.Uttam, J., Hutton, E., Coulombe, P. A., Anton-Lamprecht, I., Yu, Q. C., Gedde-Dahl, T., Jr, Fine,

J. D. and Fuchs, E. (1996). The genetic basis ofepidermolysis bullosa simplex with mottledpigmentation. Proc. Natl. Acad. Sci. USA 93, 9079-9084.Wagner, M., Hintner, H., Bauer, J. W. and Onder, K.(2012). Gene expression analysis of an epidermolysisbullosa simplex Dowling-Meara cell line by subtractivehybridization: recapitulation of cellular differentiation,migration and wound healing. Exp. Dermatol. 21, 111-117.Windoffer, R., Kolsch, A., Woll, S. and Leube, R. E.

(2006). Focal adhesions are hotspots for keratin filamentprecursor formation. J. Cell Biol. 173, 341-348.Windoffer, R., Beil, M., Magin, T. M. and Leube, R. E.

(2011). Cytoskeleton in motion: the dynamics of keratinintermediate filaments in epithelia. J. Cell Biol. 194, 669-678.Woll, S., Windoffer, R. and Leube, R. E. (2007). p38MAPK-dependent shaping of the keratin cytoskeleton incultured cells. J. Cell Biol. 177, 795-807.Yamada, S., Wirtz, D. and Coulombe, P. A. (2002).Pairwise assembly determines the intrinsic potential forself-organization and mechanical properties of keratinfilaments. Mol. Biol. Cell 13, 382-391.Zhao, Y., Gartner, U., Smith, F. J. D. and McLean,W. H. I. (2011). Statins downregulate K6a promoteractivity: a possible therapeutic avenue for pachyonychiacongenita. J. Invest. Dermatol. 131, 1045-1052.

Journal of Cell Science 125 (17)3928