Research Team:

Dr. Jun Qin's Laboratories
MCB and BMB Departments
Baylor College of Medicine
One Baylor Plaza, BCM125
Houston, TX 77030

Funding and Support:

Alkek Center for Molecular Discovery at Baylor College of Medicine
EPICOME Contributions: Mass Spectrometry Research Facilities

McLean Foundation at Baylor College of Medicine
EPICOME Contributions: Mass Spectrometry Research Facilities

National Institutes of Health (NIH)


NIH (NIDDK,NHLBI,NIEHS) U19-DK062434 NURSA CONSORTIUM:PROTEOMICS
In the next 5 years of the NURSA project, we will carry out a series of advanced data acquisitions that will provide a proteomic atlas for the "systems biology" of signal-regulated NR pathways in vivo. Such highthroughput data acquisition projects are not themselves possible as R01 mechanisms, yet the accumulated data will greatly stimulate hypothesis-driven research in the entire field of nuclear receptor science. We will pursue the isolation and identification of coregulator (CoR) complexes in mouse tissues, improving protocols For generating protein extracts from mouse tissues of liver, adipocytes, uterus and brain for proteomic analyses, and isolating and identify coregulator protein complexes from mouse tissues for analysis by mass spec. We will carry out profiling of post-translational modifications (PTM) of coregulator complexes and we will also profile coordinated PTMs of NR and CoRs following physiologic signaling.
EPICOME Contributions: msGPS CCI Resource

NIH R01-HD008188 REPRODUCTIVE HORMONES - BIOLOGICAL AND MOLECULAR ACTIONS
Steroid hormones stimulate growth, maturation and the development of new biochemical capacities in their endocrine target organs and are keys to understanding multiple diseases. They act by binding to their cognate nuclear receptors and recruiting a series of coregulator (coactivator/corepressor) proteins that carry out all substeps of transcription and also influence non-nuclear functions of the hormones. Although the basic overall pathways by which sex steroids exert their biochemical actions are known, the detailed mechanisms by which they act to regulate functions in normal and pathological tissues remain elusive. For example, in the past 12 years, we have discovered the existence of NR coactivators, determined how they function at various steps of chromatin remodeling and transcription, and realized their enormous potential in influencing the initiation and progression of disease. Now, we need to understand the mechanisms by which coactivators function in precise detail. For example, how does one coactivator perform so many different important functions in mammalian tissues? How does a coactivator become `activated' to form distinct multimeric coactivator complexes for cellular functions? How does a coactivator differentially bind to NRs (or other transcription factors) for transcriptional activation at select promoters? How does a coactivator direct distinct subreactions of transcription (eg., chromatin remodeling, transcriptional initiation, RNA chain elongation, RNA splicing, termination, etc.)? How can one coactivator participate in regulating molecular substeps of transcription on one hand, and then on the other hand, function in totally different cellular compartments to control mRNA translation, mitochondrial biology, or membrane initiated cell motility? Finally, how does the same coregulator function as a `coactivator' in certain instances, and as a `corepressor' in other instances? Even more perplexing, how can a specific coactivator function as a stimulator (eg., oncogene) in certain cell contexts and a repressor (eg., tumor suppressor) in others? We are excited to have recently discovered that the secret to these functions lies in the `posttranslational modification (PTM) coding' of coactivators. It is our belief that understanding this PTM coding, will disclose the mechanistic secrets of all normal function and the information required for the diagnosis and treatment and prevention of myriad endocrine and reproductive diseases.
EPICOME Contributions: msGPS CCI Resource, ERE Resource

NIH R01-CA084199 BIOCHEMICAL ANALYSIS OF THE BRCA1 PROTEIN COMPLEX
The long-term goal of this grant is to understand the functions of tumor suppressor BRCA1 protein. We previously purified and identified a BRCA1 protein complex, BASC. The composition of this complex has led us to propose that BASC functions as genome surveillance complex in which the DNA repair proteins function in the upstream of the DNA damage response pathway to detect DNA lesions of different types. We will further test the genome surveillance complex hypothesis. Despite the mounting evidence that BRCA1 functions in DNA damage response, the precise roles of BRCA1 and its associated partners in the conceptual framework of DNA damage response need to be addressed. We hypothesize that TopBP1 functions as an adaptor and forms a checkpoint module with BRCA1 in response to DNA damage that parallels that of scRad9 and scRad53 in S. cerevisiae. This hypothesis thus expands the effector enzymatic activity to include an E3 ligase in the DNA damage response, adding to the effector enzymes of kinases so far (Chkl and Chk2). Furthermore, we propose that BRCA1 exerts its functions through its substrates. The many implicated functions of BRCA1 that are often seemingly unrelated and confusing may now be rationalized as the effects of different substrates. The identification of BRCA1 substrates through which the checkpoint activation is executed is another goal of this proposal. The specific aims are (1) To test the hypothesis that RFC and/or BLM within the BASC function upstream in the response to DNA replication stress, (2) To purify BRCA1 complexes after DNA damage, (3) To test the hypothesis that TopBP1 and BRCA1 form a checkpoint module that parallels that of scRad9 and scRad53, and (4) To identify and characterize substrates of the BRCA1 ubiquitin ligase activity.
EPICOME Contributions: msGPS CCI Resource

NIH R01-GM080703 FUNCTIONAL ANALYSIS OF A PUTATIVE ATM/ATR SUBSTRATE RFWD3 The tumor suppressor protein p53 is a key regulator of cell cycle arrest, apoptosis and genomic stability. Mutations of p53 that compromise its function occur in 50% of human cancers. In unstressed cells, p53 is maintained at low levels; this is achieved by the ubiquitin-proteasome system, in which p53 is ubiquitinated by E3 ligases and targeted for degradation. In response to genotoxic stress, p53 needs to be rapidly stabilized to upregulate gene transcription. At least five ubiquitin ligases (E6-AP, HDM2, ARF-BP1, COP1, and PIRH2) and an ubiquitin-specific protease (HAUSP) that opposes the ligases have been identified to mediate ubiquitin- dependent proteasomal degradation of p53. However, how these ligases and protease coordinate to regulate p53 stability, particularly after DNA damage is not clear. DNA damage checkpoints are signal transduction pathways that stop or delay the cell cycle progress in the presence of DNA damage. Two kinases, ATM and ATR, are key upstream regulators of this pathway. In a proteomic screen for novel substrates for our preliminary studies, we found that ring finger and WD repeat domain 3 (RFWD3) protein is a putative ATM/ATR substrate that is required for cell cycle arrest. Furthermore, we found that RFWD3 is required for stabilization of p53 in response to DNA damage. We hypothesize that RFWD3 is a novel E3 ligase that plays an important role in mammalian DNA damage response network in part by serving as a positive regulator of p53 stability. We propose to 1) determine the mechanism by which RFWD3 regulates DNA damage checkpoint; whether this is achieved by modulating p53 ubiquitination and stability; 2) to map the RFWD3 phosphorylation sites and test how its phosphorylation by ATM/ATR regulates its function; 3) to isolate RFWD3-associated complexes in cycling cells and in response to DNA damage and identify their components by mass spectrometry; we aim to find and validate important RFWD3 regulators and its E3 ligase substrates, thus revealing clues for its other cellular functions. This is a newly identified putative ATM/ATR substrate that, when inactivated by siRNA, causes defects in p53 accumulation and cell cycle arrest. Thus, RFWD3 may play an important role in the mammalian DNA damage response network by serving as a novel E3 ligase that positively regulates p53 stability. Uncovering new positive regulator of p53 and understand its function will potentially advance our knowledge of cancer development and treatment.
EPICOME Contributions: msGPS CCI Resource