Zhou Songyang, Ph.D.
Biochemistry & Molecular Biology, Pharmacology
- Ph.D., 1995, Molecular and Cellular Physiology, Tufts University
- Postdoctoral, Biology, 1996-1998, MIT
- 1995-1996, Harvard University,
- 1996-98, Irvington Institute Fellowship
Molecular Mechanisms of Signal Transduction
Our laboratory is interested in studying the molecular mechanisms of signaling pathways that regulate cell survival and aging through functional genomic approaches. The systems that we use to study these questions are also uniquely suited for studying stem cell biology. Through these model systems we hope to address how survival factors control stem cell survival and self-renewal, and the role of telomere associated factors in both somatic and stem cell biology.
Genetic Screens, Proteomic and Functional Genomics
(1) Genome stability and telomere maintenance
In the past few years, one major focus of our laboratory has been to elucidate the protein complexes that participate in regulating genome stability, particularly those involved in telomere end protection, maintenance, and signaling. Towards this end, we have been systematically analyzing protein networks that regulate human telomeres. During this process, we identified a novel telomere binding protein PTOP/TPP1 that interacts with telomeric proteins TIN2 and POT1, and participates in telomere regulation. Interestingly, we found that the TPP1 and POT1 acts as a heterodimer to bind to telomere DNA overhangs with higher affinity, and interacts directly with telomerase. We have also discovered that the six core telomere proteins (TRF1, TRF2, POT1, RAP1, TIN2, and TPP1) can form a high-order complex, the telosome. To date, we have purified more than 20 protein complexes related to telomere maintenance in human cancer cell lines. Analyses of these complexes have led us to propose the Telomere Interactome, an interaction network of telomere regulators in mammalian cells. Our analysis indicates that diverse types of enzymes and activities are present at the human telomeres. These activities and factors are recruited and assembled on the telomeres through the telosome platform and its sub-complexes. We are interested in the mechanisms by telomeric proteins regulate diverse signaling pathways.
(2) ERM-mediated genetic screens
Genetic screens play an important role in identifying central genes that regulate various signaling pathways. My laboratory has developed several tools for gain-of-function genetic screens in mammalian cells. One of these is a novel retrovirus-based technique (Enhanced Retroviral Mutagen: ERM) for random gene tagging and manipulation. ERM takes advantage of the splicing machinery to generate N-terminus tagged endogenous proteins in cells, does not require construction of cDNA expression libraries, and allows genome-wide inducible activation of cellular genes. Depending on the tag sequences engineered in ERM, we can generate a variety of in vivo expression libraries to dissect signaling pathways. For example, we have performed ERM screens to identify genome-wide mammalian genes that regulate cell survival and self-renewal. In addition, we have used fluorescent protein (such as GFP and YFP) ERM tags to determine genome-wide the subcellular localizations and interaction networks of human proteins.
(3) High-throughput analysis of PTM-dependent protein-protein interactions
Most signal proteins contain homology sequences (domains) that are necessary for specific protein-protein interactions. These domains ensure proper targeting, assembly and firing of protein signal complexes. The identification of the targets of these domains therefore represents a critical step towards the understanding of signal networks.
In vitro interaction analysis using OPAL. We have taken a unique approach to systematically study the binding specificities of protein domains, by developing an Oriented Peptide Array Library (OPAL) technique to determine the specificities of a variety of important signaling domains. Through OPAL, we are able to examine protein-protein interaction in vitro in a high throughput manner. Given the recent exciting findings on post-translational modifications (PTM) such as protein phosphorylation, acetylation, and methylation, we envision that OPAL will be an extremely powerful tool for probing the molecular basis for these types of interactions.
Cellular protein-protein interaction analysis. To study protein-protein interactions in vivo, several improved methods have been developed. Many of these methods utilize protein fragment complementation to detect protein-protein interactions. For example, the biomolecule fluorescence protein complementation (BiFC/PCA) approach was developed to detect protein-protein interaction in live cells. My lab has been actively modifying and employing these methods to study protein-protein interactions in cultured cells. Furthermore, we combined ERM and BiFC, two powerful techniques, to develop the RePCA method for the isolation of direct interacting proteins of master signaling regulators.
(4) Embryonic stem cell proteomics and functional studies
Telomere maintenance is essential for the integrity of mammalian cells and tightly linked to cell senescence and aging. Telomere associated factors and telomerase have also been implicated in regulating stem cell function and fate. For example, mice defective in PTOP/TPP1 expression (acd mice) also have defects in stem cells and germ cells. Recent studies also suggest that inactivation or overexpression of telomerase can result in abnormal stem cell phenotypes. To understand the role of telomere proteins in stem cell function, we will use proteomic approaches to isolate telomeric protein complexes from stem cells. In addition, we will utilize available proteomic and genetic tools to define the function of telomeric proteins and self-renewal transcription factors in stem cells.