Paul Matsudaira

WI/MIT BioImaging Center, Dept Biology and Div Biological Engineering, MIT

Cellular machinery, biomechanics, and bioinformatics

IC-21 macrophage cells transfected with GFP, imaged in 3D, and rendered as a projected solid (Imaris)

•bioengineering models describe biological processes•complex movements via cellular machines•mechanics + chemistry•structures capture states of movement•informatics/computing resources

ManipulateMolecular GeneticsChemical GeneticsCell Engineering

MeasureBiochemistry

ImagingProteomics

Bioelectronics

Systematic Experiments

ModelDifferential Eq. Matrices

Bayesian NetworksBoolean Networks

MineBioinformatics

GenomicsData Semantics

Quantitative Models

Quantitative Biology Paradigm

courtesy of CSBi

Cell biologists study large, dynamic structures

G. Borisy and S. Tatiana

1 nm 1 µm 1 mm

cellsorganelles

molecules protein machines

NMR, x-rays

cryo electron microscopy

single particle det. dynamic 4D light microscopy

resolution and size scale

Overlapped structural and cell biology approaches

zone of adhesion and extenstion

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Forces generate movements, machines generate forces

fimbrin-EGFPtimelapse 30 sec

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Changes in structure-assembly, force, chemistry

mother adhesiondaughter adhesions

top side

Formation of cell adhesions at the leading edge of a macrophage cell(J. Evans et al J. Cell Biol. in press)

•multispectral fluorescence data acquisition•video rate data acquisition•cellular tomography and single particle cryoEM imaging•high content screening•whole proteome studies

Cell Biology Appetite for More Information

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High content: Tracking all

cell adhesions

(J. Evans et al J. Cell Biol. in press)

Dynamics of structure modulated by microtubules

10 µm paclitaxel

10 µm demecolcine

(J. Evans et al J. Cell Biol. in press)

Imaging storage/processing requirements

object file sizegenome sequence 12 MBprotein structures1 2 GB2D localization2 6 GB3D localization3 300 GB4D localization4 3.6 TB152kDa (ave. MW) x 110 MW/residues x 10 atoms2512x512 two channel, 16 bit TIFF350 image slices/stack412 images/series

The BioImaging Pipeline

acquisition

management

processinganalysis modelling

Podosomes split, merge, and appear de novo

polar assembly

simple

dendroid

genomics imagingtraces counts

ATGC voxelsequence image

function function

Imaging is an informatics science

WI/MIT BioImaging Net

terminal

terminal

terminal

terminal

Origin3400

4 TB

confocalmicroscopy

Origin2400

4 TB

IBM Power4 6550

Cryo-EM

2-photon microscopy

Tape

RAID30TB

imaging modes

Expansion and management of imaging data

channel 0 channel 1 channel 2

multi-spectral (channels) 12 or 16-bit acquisition

256 x 256 1 130 KB3 910 KB

1024x 1024 1 2 MB3 14 MB

image size (pixels) channels file size

1+2+3

(x,y,z,t,ch) file size 256 x 256 x 1 x 3 400 KB 256 x 256 x 50 x 3 20 MB 256 x 256 x 50 x 20 x 3 400 MB 1024 x 1024 x 50 x 3 800 MB 1024 x 1024 x 50 x 20 x 3 16 GB

3D/4D image data (stacks)

35KB to 8.2 MB per channel/stack (X:Y:Z or X:Y:T)

NMR

x-raycryoEM

light

microscopy

raw data12 or 16-bit

deconvolved data

32-bit

intermediates32-bit

raw data32-bit

40 MB 80 MB 80 MB 80 MB 80 MB

40 MB 120 MB 200 MB 280 MB 360 MB

accumulated file size

Image processing (deconvolution) RAM 256 x 256 x 50 x 3 (x,y,z,ch)

3.2 GB

ch1 vs. ch2ch1 vs. ch3 ch2 vs. ch3ch1 vs ch2 vs. ch3

colocalization

Total data accumulation

rendering

1 GB

movies

object analyses

1 GB

45 5 335 45 5 335 45 5 335 45 5 335 45 5 3355 5 53 5 5 53 5 5 53 5 5 53 5 5 5345 43 53 45 43 53 45 43 53 45 43 53 45 43 53

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45 5 335 45 5 335 45 5 335 45 5 335 45 5 335

5 5 53 5 5 53 5 5 53 5 5 53 5 5 5345 43 53 45 43 53 45 43 53 45 43 53 45 43 53

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5 5 53 5 5 53 5 5 53 5 5 53 5 5 5345 43 53 45 43 53 45 43 53 45 43 53 45 43 53

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5 5 53 5 5 53 5 5 53 5 5 53 5 5 5345 43 53 45 43 53 45 43 53 45 43 53 45 43 53

545578 8787 8786 545578 8787 8786 545578 8787 8786 545578 8787 8786 545578 8787 878645 5 335 45 5 335 45 5 335 45 5 335 45 5 335

5 5 53 5 5 53 5 5 53 5 5 53 5 5 5345 43 53 45 43 53 45 43 53 45 43 53 45 43 53

545578 8787 8786 545578 8787 8786 545578 8787 8786 545578 8787 8786 545578 8787 878645 5 335 45 5 335 45 5 335 45 5 335 45 5 335

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1 GB ?

raw 0.4 GBpost-deconvol 3.6 GBanalysis 7.2 GB

total 11.2 GB

terminal

terminal

terminal

terminal

SGI Origin3400

32 GB RAM4 TB

Zeiss510 META ftp

data acquisition data storageimage restorationimage analysisbatch rendering

database management remote visualizationrenderingbatch design

Current WI (local) imaging net

stack (512 x 512 x 50 x 12 x 6)4.8 GB

Microtubules at podosomes

tubulinactinfimbrin

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Dynamic structures from light microscopy

fimbrin-EGFPtimelapse 15 sec

side/bottom view top view

‘finger’

‘palm’‘finger’

‘finger-tip’‘palm’

assembly of cell adhesions in a live cell

Dynamic Cell Adhesions - Static