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ISMB/ECCB
ISMB/ECCB Stockholm 2009: ISMB/ECCB
Keynote: Mathias Uhlen - A global view on protein expression based on the Human Protein Atlas
July 1, 2009 - Comment - Share
Introduction: Works a lot on affinity reagents. Invented and developed pyrosequencing technology (http://en.wikipedia.org/wiki...) now used in 454 - Allyson Lister
Out line of the talk - 1. systematic biology -introduction, 2. HPR project 3. The Human protein Atlas - Venkata P. Satagopam
18th century - biologist. 19th - chemist (1/3 of elements discovered in Sweden in this century). 20th - physicists and at the end, computer scientist. He'd now like to say that the 21st century is the century of medicine. - Allyson Lister
HPR one of the largest projects in Sweden wrt funding, about 100 million euro so far - Oliver Hofmann
An impressive log-scale plot of number of bases sequenced since 1965. - Allyson Lister
developer of sequencing by synthesis via pyrosequencing in late 90s -- basis of 454 technology - Andrew Su
Personalized genomics ... 454 technology developed in our lab - Venkata P. Satagopam
Bioinformatics is the key in the new era of genomics. - Allyson Lister
95% of drugs (still) aimed at proteins - Oliver Hofmann
(Personal opinion: I like how it's not the "post-genomic era", but a new era of genomics :) ) - Allyson Lister
95% of drugs today target proteins. Thus, studying proteins is studying for the future - Diego M. Riaño-Pachón
Systems biology /omics is going to be fantastic in the next 10 years. - Allyson Lister
(That, or we find better ways of interfering with RNA) - Oliver Hofmann
Image of contradictory sign in Paris: you know where you want to go, but not how to get there. - Allyson Lister
We know where we want to go (characterize all proteins), but not sure how to get there (due to a lack of high-throughput methods) - Oliver Hofmann
The generation game - Nature july 7, 2007 - Venkata P. Satagopam
antibodies are the core tool for probing proteins - Andrew Su
but they are too cross-reactive - Diego M. Riaño-Pachón
Human antibody initiative HAI - Venkata P. Satagopam
human anitobdy initiative (HAI) -- uhlen, M. Synyder, P. Hudson -- generate comprehensive and validated antibody collection - Andrew Su
validation of commercially available antibodies - Venkata P. Satagopam
average success rate of commercial antibodies is 49% - Andrew Su
some companies 100% antibodies works fine, some companies 0%, in general 50% works fine - Venkata P. Satagopam
Antibodypedia -- a portal for validated anitbodies (we need to add a link from Gene Wiki...) - Andrew Su
From the website: "The antibodypedia is a community-based portal showing application-specific validation of publicly available antibodies to human protein targets. Each protein binder (antibody or other affinity reagent) has been scored in an application-specific manner into three main categories (supportive, uncertain and non-supportive)" - Oliver Hofmann
@Oliver - nice! - Allyson Lister
If you have 2 antibodies, you can compare results in various assay platforms so he wants to develop paired antibodies for every protein target. - Allyson Lister
Nat Methods 2008: High-througput method to identify epitopes - Oliver Hofmann
6 months ago published a paper Nature methods (december 2008) - Venkata P. Satagopam
(bummer, antibodypedia doesn't use mediawiki so can't assess current usage...) - Andrew Su
(I am entirely too short-sighted to read the author lists half of the time. Sigh) - Oliver Hofmann
HPR - The human proteome resource - Venkata P. Satagopam
(grumble grumble, antibodypedia creates YAI -- yet another identifier) - Andrew Su
(++ HPA -- uses ensembl gene IDs...) - Andrew Su
hpr is a multi-disciplinary program - Venkata P. Satagopam
(Proteinatlas seems to be using ENSG/ENSP/UniProt) - Oliver Hofmann
The gene factory does about 200 clones per week, and is in full production. - Allyson Lister
(HPA ids- mapped to uniprot also) - Venkata P. Satagopam
Close to 34.000 clones in the database - Oliver Hofmann
200 clones per week, 33,925 clones total (all human?) - Andrew Su
(@Andrew - indeed) - Allyson Lister
Open source (but in-house?) LIMS developed - Oliver Hofmann
(would be interesting to compare to origene collection of mammlian clones) - Andrew Su
The antigen design uses PRESTIGE, which is a bioinformatics approach to select antigens using the protein epitope signature tag (PrEST). - Allyson Lister
antigen design -- used PRESTIGE a bioinformatics approach to select antigen for antibody - Venkata P. Satagopam
read of this project is protein expression profiling - Venkata P. Satagopam
readouts -- immunohistochemistry (IHC) and IF (immunofluorescence) - Andrew Su
Organ, tissue, cellular and sub-cellular expression profiing on a protein basis - Oliver Hofmann
apply antibodies to tissue arrays (cancer focus, I think) - Andrew Su
(140+ human samples, around 200 tissues.. did someone catch the numbers?) - Oliver Hofmann
(Faq from the website: spatial distribution of proteins in 48 different normal tissues and 20 different cancer types as well as 47 different human cell line) - Oliver Hofmann
annotation of images taking place in Mumbai in India - Venkata P. Satagopam
image annotation -- difficult problem. automated anaylsis would be good, but now using indian pathologists for manual annotation - Andrew Su
$60 / 500 images for annotation? - Andrew Su
Confocal microscopy for subcellular localization, difficult to scale up to high-throughput - Oliver Hofmann
high-throughput subcellular localization in A0431 (squamous cell carcinoma), U-251MG (glioma), ??? - Andrew Su
(I think Bob Murphy talked on a similar project to map subcellular localization of proteins en masse...) (Oh, looks like it's a collaboration between the two...) - Andrew Su
They have a SVM that seems to be able to annotate 28 different parts of the cell. - Allyson Lister
2TB data each week (courtesy of 50.000 images in the same time) - Oliver Hofmann
2/3 of data come from in house data and 1/3 comes from different companies - Venkata P. Satagopam
(I wonder how the protein expression compares with our gene expression atlas http://www.ncbi.nlm.nih.gov/pubmed...) - Andrew Su
About 33% of the sample space done (6850 genes) - Oliver Hofmann
progress - started in 2005 , last week released version 5. 8,832 antibodies, covering 1/3 genes in uniprot - Venkata P. Satagopam
(And I suppose there's always more to do.. check for splice variants, truncated versions...) - Oliver Hofmann
Most recent release: 7 mln images. - Allyson Lister
(@Andrew: if there is overlap in the cell lines that could be an easy correlation analysis) - Oliver Hofmann
The next 5 yrs are also about getting the paired antibodies mentioned earlier. - Allyson Lister
all antibodies available to the public - Venkata P. Satagopam
(good good, HPA is already BioGPS plugin.. http://biogps.gnf.org/#goto=p... </shameless_plug>) - Andrew Su
central questions in proteomics - Venkata P. Satagopam
(Rodent atlas seems a bit redundant to Allen Brain Atlas? -- Oh, ABA is via in situ / RNA, this is protein. Again, would be interesting to compare...) - Andrew Su
how many proteins are expressed in a given cell? - Venkata P. Satagopam
how many protein are tissue specific? - Venkata P. Satagopam
@Oliver, probably not directly comparable by exact cell lines, but might be worth comparing by parental tissue. Need to wait until they allow downloading of data though... - Andrew Su
Ensembl "thinks" that the genes are up to 23,000, but UniProt "thinks" 20,000, but the number is probably with that (for genes coding for proteins). ("thinks" in scare quotes, as databases don't think - yet) - Allyson Lister
the size of human membrane proteome .. 5,514 human membrane proteins; covering 26% of protein-encoded genes - Venkata P. Satagopam
proteins expressed in normal cells - 6,800 antibodies towards (>25% of all protein encoding genes). 65 normal cell types (from 45 different tissue types) - Venkata P. Satagopam
70% of proteins expressed in a given cell, approx even distribution across # of cell lines (not what we observed on gene expression data, which had distinct peaks at tissue-specfiic and ubiquitous) - Andrew Su
80% of proteins expressed on average in cell lines (surprisingly high to me...) - Andrew Su
(@Andrew: protein selection might be biased towards the ones that are well expressed / had known anitbodies / ...) - Oliver Hofmann
9% of proteins cell type specific, 62% expressed across 3 different cell types - Oliver Hofmann
ubiquitous expression, but differing levels - Andrew Su
(interesting cytoscape visualizations of cell type / tissue specificity) - Andrew Su
In the Atlas: < 2% specific to a single cell type (84 proteins), well known ones like insulin - Oliver Hofmann
Includes a number of uncharacterized proteins with no known function - Oliver Hofmann
PROSPECTS: PROteomics SPECification in Time and Space - Allyson Lister
"Complementary technologies, including mass spectrometry, cryoelectron microscopy and cell imaging will be applied in innovative ways to capture transient protein complexes and the spatial and temporal dimensions of entire proteomes." - Oliver Hofmann
MCF-7 data with IHC, Mass spec - Oliver Hofmann
next generation seq of cDNAs from U2-OS human cell line 76% detected by mRNA seq - Venkata P. Satagopam
76% genes detected by next gen mRNA sequencing in U2-OS - Andrew Su
again -- mostly ubiquitous expression (now on mRNA level), but differing levels - Andrew Su
high fractions of all proteins expressed in human cells, tissues and organs - Venkata P. Satagopam
Lack of specificity not good knows for those looking for good antibody targets for therapeutic purposes - Oliver Hofmann
the quantity of proteins, rather than their presence /absence, is the key to cell identity - Diego M. Riaño-Pachón
few cell-specific proteins (<1%) and group-specific proteins (<10%) - Venkata P. Satagopam
find biomarkers for early detection of disease ... it is very good for human mankind - Venkata P. Satagopam
(Proteome 2008) Suspension bead arrays - Oliver Hofmann
mg/ml to pg/ml range (dynamic range of protein concentration in blood 10^12 ) - Oliver Hofmann
working on kidney disease in collaboration with astrazanica .... for detection of biomarkers - Venkata P. Satagopam
Developing 'next generation' plasma profiling, scale to one million assays / months - Oliver Hofmann
(has he mentioned availability of these antibodies? Commercially available? antibody-producing lines?) - Andrew Su
They're part of ENGAGE. - Allyson Lister
commercially available @Andrew - Allyson Lister
first draft of the human proteome by 2014 - Venkata P. Satagopam
Aim to have the draft version of the human proteome by... see above - Oliver Hofmann
(@Andrew I think via Prestige Antibodies (I remember the cute advertising slide). Does that mean advertising works?) - Allyson Lister
Nature, "The big ome" - 24 April 2008, editorial - Allyson Lister
tissue-specificity is achieved by precise regulation of protein levels in space and time - Venkata P. Satagopam
Prestige Antibodies through Sigma: http://www.sigmaaldrich.com/life-sc... - Andrew Su
science 26 sep 2008, vol 321 pages 1758-1761 - Venkata P. Satagopam
"Proteomics Ponders Prime Time", Science, 26 September 2008, in response to the Nature article - Allyson Lister
new lab in Stockholm coming soon, Science for Life Laboratory - Venkata P. Satagopam
New Science for Life laboratory being established, see http://www.newsdesk.se/pressro... - Oliver Hofmann
Q: importance of splice isoforms -- A: complexity that is currently not considered due to technical complexity (to be saved for second phase) - Andrew Su
Q: conclusions on tissue specificity have bias based on antibody availability? A: bias of commercial antibodies possible, but only 1/3 of data. Data they are generating based on walking down chromosomes (I think?), so don't expect bias... Also, some of ubiquitous expression is due to cross-reactivity. (first mention of this...) - Andrew Su
Q: perspective for gene therapy or antisense therapy, more generally non-protein based therapies. A: pharma shifting from small molecules to biologics (not sure about a "shift" rather than "expansion"). Gene therapy problem is getting into all relevant cells. Ubiquitous expression of proteins the root cause of side effects for protein-based targets, possibly... - Andrew Su
(only to get the blog on top of the ISCB portal site; the figures messed up our layout) - Reinhard Schneider