Fate of mRNA after RNA processing events in nucleus

Hafidh Adv Exp Med Biol. 722:118, 2011

“Proofreading and spellchecking: A two-tier strategy for pre-mRNA splicing quality control”

Egecioglu & Chanfreau RNA 17: 383, 2011

DEAD-box helicases (ATPase Prp proteins) unwind incorrect base-pairing

What happens if splicing pathway is defective?

Egecioglu & Chanfreau RNA 17: 383, 2011

RNA degradation pathways for RNAs that escaped splicing or have splicing mistakes

“Quality control” removal of defective transcripts

Nonsense-mediated mRNA decay (NMD)

- surveillance system to remove defective RNAs

- mRNAs with premature stop codons are rapidly degraded

Gonzalez Gene 274: 15, 2001

Observations:

- in “pioneer round” of translation, if EJC (exon joining complex) not removed by ribosomal machinery, it acts as tag for RNA degradation pathway

Wilusz Genes & Dev 15:2781, 2001

Steady state levels of RNase-L mRNA quantified by qPCR and normalized relative to constitutively expressed rpl13a mRNA

3’UTR deletion constructs

Example of AREs (AU-rich cis-elements) in 3’ UTR of RNase-L mRNA

Deletion constructs were transfected into 293T cells and RNase-L expression was monitored by Western blots

Li et al. J Biol Chem 282:7950, 2007

Interpretation of data? What experiments might be interesting to do next?

Cis-determinants of mRNA stability

ZC5+

Kloc Cell 108: 533, 2002

Drosophila egg: nanos mRNA

Xenopus oocyte: fatVg mRNA

Chicken fibroblast: -actinmRNA (red) in leading lamellae

Mechanisms that target certain mRNAs to appropriate subcellular site?

Heredia Curr Opin Cell Biol 16:80, 2004

Localization of mRNAs within eukaryotic cells

Becalska & Gavis Develop 136:2493, 2009

“Lighting up mRNA localization in Drosophila oogenesis”

Advantages of mRNA subcellular localization (and localized translation)?

Martin & Ephrussi Cell 136:719, 2009

- mRNAs (eg nanos, bicoid, oscar, K10…) important for developmental processes are synthesized in nurse cells and move to oocyte along microtubules

mRNA localization in Drosophila oocyte / embryo

Saxton Cell 107:707, 2001

Kloc & Etkin J.Cell Sci. 118:269, 2005 (Fig.4)

Serano Develop. 121: 3809, 1995

44 nt TLS (transport/localization sequence) with stem-loop structure in 3’UTR

Model for Drosophila K10 mRNA movement?

KSD

Serano Develop. 121: 3809, 1995

Mutational analysis of stem-loopin 3’ UTR of K10 mRNA

Huang Curr Opin Cell Biol 16:308, 2004

Mechanisms for localized translation of mRNAs