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Our laboratory seeks to elucidate the pathogenetic mechanisms underlying neurodegenerative disorders. We are interested in understanding how genetic mutations affect the expression and function of genes important for cell death and critical for the pathogenesis of neurodegenerative diseases. The lab is focusing on two areas of research: the role of pre-mRNA splicing regulation in neurodegeneration and the molecular mechanisms modulating cell migration.
Pre-mRNA splicing is a crucial step for gene expression because the vast majority of human genes contain one or more intervening sequences (introns) that must be accurately removed to form the mature and functional messenger RNAs (mRNAs). Alternative splicing is a major mechanism for regulating mammalian gene expression and for generating the complexity of human proteomes. Mutations that affect pre-mRNA splicing cause a large number of diseases. Alternative splicing regulates the expression and function of programmed cell death (PCD) genes. A number of PCD genes, including Bcl-x, ced4/APAF1 and caspases, produce functionally antagonistic products by undergoing alternative splicing. We have established a model using the caspase-2 (casp-2) gene. An intronic regulatory element has been identified to control the balance between anti-apoptotic and pro-apoptotic isoforms of casp-2 gene products. Similar intronic elements are present in other human caspase genes in the regions critical for their enzymatic activities. These intronic elements may play a role in regulating the formation of functionally antagonistic caspase gene products. Our work has provided direct evidence that splicing factors can regulate casp-2 alternative splicing and has suggested that alternative splicing may be an important regulatory mechanism for PCD. We are using molecular, biological and biochemical approaches to dissecting cis-elements and trans-factors critical for alternative splicing regulation of caspase genes. We are testing whether cell death signals such as chemotherapeutic reagents trigger changes in the expression or function of splicing regulators. This work will not only further our understanding of PCD regulation but also provide insights for designing new therapies for diseases associated with excessive or insufficient cell death, including inflammation and cancer.
Accumulating evidence supports the involvement of pre-mRNA splicing defects in pathogenesis of neurodegenerative disorders such as dementia and retinal degeneration. Aberrant splicing of the human tau gene leads to tauopathy, including fronto-temporal lobe dementia (FTD) and progressive supranuclear palsy (PSP). A large number of splicing mutations have been identified in human tau gene in FTD patients. We have initiated biochemical and molecular studies to identify cis-elements and trans-factors important for tau alternative splicing regulation.
Genetic defects in ubiquitously expressed splicing factors lead to neuron-specific diseases, including autosomal dominant retinitis pigmentosa (adRP) and spinal muscular atrophy (SMA). We have established biochemical and cell biological models to examine pathogenetic mechanisms underlying retinal degeneration caused by such splicing defects. More specifically, we are investigating how mutations in PRPF31 and survival of motor neuron (SMN) genes cause RP and SMA respectively. Our recent experiments have identified functional links between different adRP genes. Our studies have begun to reveal how defects in general splicing factors lead to neuron-specific manifestations.
Cell migration is essential for the development of multi-cellular organisms. Chemotactic cell migration plays a crucial role in both physiological and pathological processes. Using primary culture systems, we have identified a family of secreted proteins, Slit, that regulate neuronal migration and modulate leukocyte chemotaxis. We demonstrated that Slit proteins are ligands for the transmembrane protein Roundabout ( .Robo) and that Slit-Robo signaling in neuronal migration is transduced by a novel family of slit-robo-regulated GTPase activating proteins (srGAPs) and Rho GTPases. Our studies also suggest a conservation of guidance mechanisms for different cell types. More recently, we developed a fluorescence-resonance-energy-transfer (FRET)-based approach that allows us to visualize the dynamic changes in the activity of Neuronal Wiskott-Aldrich Syndrome protein (N-WASP) in living cells. Using time-lapse microscopy, we are characterizing the temporal and spatial activation of N-WASP in migrating cells in response to guidance cues. The ability to directly visualize the dynamic regulation of WASP activities in living cells at high spatial and temporal resolutions provides a powerful tool for studying a number of distinct biological problems, including growth-factor signaling, vesicle trafficking, axon guidance and directed cell migration. Combining these molecular, biochemical and cell biological approaches will help elucidate molecular mechanisms regulating cell migration.
Publications:
Li H-S, Chen JH, Wu W, Fagaly T, Zhou L, Dupuis S, Jiang ZH, Nash W, Gick C, *Wu JY, and Rao Y (1999). Vertebrate Slit, a secreted ligand for the transmembrane protein Roundabout, is a repellent for olfactory bulb axons. Cell 96:807-818. [* co-corresponding author]
Wu W, Wong K, Chen JH, Jiang ZH, Dupuis S, *Wu JY, and Rao Y (1999). Directional guidance of neuronal migration in the olfactory system by the secreted protein Slit. Nature 400:331-326. [* co-corresponding author]
Jiang ZH, Cote J, Kwon JM, Goate AM, and Wu JY . (2000) Aberrant splicing of tau pre-mRNAs caused by intronic mutations associated with the inherited dementia FTDP-17. Mol. Cell. Biol. 20:4036-4048
Cote J, Dupuis S, Jiang ZH and Wu JY. (2001a) Caspase-2 alternative splicing: identification of a novel intronic element acting as a decoy acceptor site. Proc. Natl. Acad. Sci. USA 98:938-943
Wu JY*., Feng, L., Park, H.-T., Havlioglu, N., Wen, L., Bacon, K. B, Jiang, Z., Tang, H., Zhang, X.-C., and Rao, Y. (2001). The neuronal repellent Slit also inhibits leukocyte chemotaxis induced by chemotactic factors. Nature 410:948-952 [* co-corresponding author]
Jiang Z, Tang H, Havlioglu, N. Zhang, X, Stamm S, Yan R and Wu JY. (2003) Mutations in tau exon10 associated with FTDP-17 alter the activity of an exonic splicing enhancer to interact with Tra2beta. J. Biol. Chem. 278(21):18997-9007.
Liu G, Beggs H, Jürgensen J, Park, H.-T, Tang, H, Gorski J, Jones KR, Reichardt LF, Wu JY & Rao Y. (2004) Netrin Requires the Focal Adhesion Kinase and the Src Family Kinases to Induce Axon Outgrowth and to Attract Axons. Nature Neuroscience 7:1222-1232
Yuan L, Kawada M, Havlioglu N, Tang H and Wu JY. (2005) Mutations in human PRPF31 gene cause splicing defects in rhodopsin, a retina-specific gene: a potential mechanism for retinal degeneration. J. Neurosci 25:748-757 (as a highlighted paper)
