Molecular Systems Biology 6; Article number 406; doi:10.1038/msb.2010.63
Citation: Molecular Systems Biology 6:406& 2010 EMBO and Macmillan Publishers Limited All rights reserved 1744-4292/10
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NEWS AND VIEWS

Transcriptional regulation is only half the story
Joshua B Plotkin*
Department of Biology, University of Pennsylvania, Philadelphia, PA, USA* Corresponding author. Department of Biology, University of Pennsylvania, 433 S. University Avenue, Philadelphia, PA 19104, USA.
Tel./Fax: þ 1 617 571 9341; E-mail: jplotkin@sas.upenn.edu

Molecular Systems Biology 6: 406; published online 24 August 2010; doi:10.1038/msb.2010.63
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The central dogma of molecular biology, articulated by FrancisCrick, posits a flow of information from DNA to RNA toprotein. Although the Human Genome Project hashelped elucidate the first step in this cascade, the relationshipbetween mRNA abundance and protein abundance has
resisted systematic quantification, especially in higher eukaryotes. In their recent publication in Molecular Systems Biology,
Vogel et al (2010) use a combination of microarrays andshotgun proteomics to quantify absolute mRNA and protein
levels for over 1000 genes in a human cell line. Their
analysis identifies sequence features related to translationand protein degradation that are as important as transcription
in determining steady-state protein levels. This work provides
an unprecedented, system-wide accounting of how information stored in our DNA determines the eventual state ofour cells.
Molecular biologists have traditionally focused on transcriptional regulation as the main determinant of protein levels
and, thus, cellular function. This focus is due, in part, to thehistorical sequence of discoveries following the work ofJacques Monod, and, more recently, to the development of
microarray- (Schena et al, 1995) and sequencing-based
technologies for large-scale mRNA quantification. The detailed
view of transcription that has emerged from high-throughput
mRNA measurements, including the inference of global
regulatory networks (e.g. Lee et al, 2002), has shaped ourunderstanding of how a healthy cell works, and also howpathologies arise and might be remedied. In fact, our knowledge of transcriptional regulation has improved so rapidly
over the past decade that it is easy to forget the diverse array of
post-transcriptional processes—including mRNA processing
and modification, miRNA modulation, translation initiation,elongation, termination, and protein degradation—that also
influence steady-state protein levels.
Not so fast, say Vogel et al (2010). Systematic studies of posttranscriptional processes have come of age, owing again to thedevelopment of technologies for high-throughput measurements. In this study, the authors used microarrays to quantify
mRNA levels, together with a sophisticated mass-spectrometry-based proteomics method called APEX (Lu et al, 2007)
to quantify soluble protein levels in a tumor cell line. The APEX
method, originally developed in yeast and Escherichia coli, is& 2010 EMBO and Macmillan Publishers Limited

the work-horse behind the present study. Under this protocol,proteins are digested into peptides, which are separated byliquid chromatography, and then ionized and sequenced withtandem mass spectrometry. In principle, protein amounts arethen quantified simply by counting the numbers of corresponding peptides observed in repeat runs. In practice, theAPEX method critically corrects for factors, such as efficiencyof ionization, that influence the a priori probability of peptidedetection. As a result, APEX provides reliable quantification ofprotein levels over five orders of magnitude.
Vogel et al. analyzed about 200 sequence features aspotential determinants of the steady-state protein levels theymeasured. The correlates considered include features such ascoding-sequence length, amino-acid composition, predicted
mRNA structure, putative miRNA target sites, and thepresence of upstream start codons. The authors observed alognormal distribution of protein-per-mRNA ratios—suggesting that many impendent factors together contrive todetermine translational efficiency and protein degradation
rates. Some of the strongest individual correlates of protein
abundance identified in the study are unsurprising: longer
coding sequences typically produced less protein, controllingfor mRNA levels, consistent with the idea that long transcripts
are translated inefficiently and are prone to protein misfolding.
Similarly, amino-acid content is also correlated with protein
abundance, controlling for mRNA levels, consistent withvariable costs associated with the depletion of different aminoacids and different propensities for protein misfolding as afunction of amino-acid composition. Furthermore, strong 50
mRNA secondary structure or the presence of upstream startcodons both reduced protein levels, again controlling formRNA. However, several features had a surprisingly smallrole: codon adaption and miRNA target sites did not
significantly influence protein abundance. The most important
take-home message, furnished by a non-linear multiple
regression, is that features related to post-transcriptional
processes, especially those found in the coding sequence,
together explained as much variation in protein levels as
mRNA levels themselves did (Figure 1). Thus, transcriptional
regulation is only half the story.
Aside from generating the largest dataset to date ofprotein and mRNA concentrations in human cells, this study
Molecular Systems Biology 2010 1

News and Views
JB Plotkin

DNA27%

Transcription

mRNAmRNA degradationof the system-wide approach to studying basic questions inmolecular biology. Without doubt, the growing literature
based on high-throughput mass spectroscopy will continue toinform our understanding of post-transcriptional regulation,much as microarrays revolutionized our understanding oftranscriptional regulation. Such measurements performed inrelatively natural cellular conditions on endogenous genes will
nicely complement manipulative experiments that interrogate
protein production using synthetic, heterologous gene constructs (e.g. Voges et al, 2004). Together, these systematic
approaches promise to elucidate the operational details ofCrick’s central dogma.

Conflict of interest
The author declares that he has no conflict of interest.

Translation

References

40%

ProteinProtein degradation

Figure 1 The relative influence of transcriptional and translational regulation onprotein abundance. Processes related to translation and protein degradation canexplain about 40% of the variation in human protein abundance, similar to theapproximately 27% explained by mRNA abundance. Together these factors canexplain two-thirds of the variation in human protein abundance.

systematically quantifies the importance of translation andprotein degradation regulatory processes, both individuallyand in aggregate. This work extends similar analyses performed
in bacteria (Nie et al, 2006) and yeast (Brockmann et al, 2007;
Wu et al, 2008), and it is preferable to analyses that are basedon mRNA and protein measurements obtained from separateexperiments. Nonetheless, this study is still limited toabout 1000 soluble proteins, measured in an asynchronous,
log-phase population of a tumor cell line, which contains
chromosomal and methylation irregularities. Moreover, thestrict separation of sequence features into those that determine
steady-state mRNA levels and those that act post-transcriptionally is problematic: some nominally post-transcriptionalfeatures, such as those that influence ribosomal initiation, may
feed back to influence steady-state mRNA levels as well (Iostand Dreyfus, 1995). Nonetheless, future studies in multiple
cell lines, ideally including membrane proteins and synchronized populations, should elucidate how protein levels differ
between and, indeed, define alternative cellular states. Suchstudies will be especially powerful when combined with highthroughput techniques for measuring ribosomal occupancy(Ingolia et al, 2009), allowing us to compare protein levels
with direct estimates of translational efficiency, and toquantify protein stabilities as well.
The quantification and analysis of protein levels for 1000
human genes is a remarkable technical feat and is emblematic
2 Molecular Systems Biology 2010

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