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Lab Members

Hua Lou
Associate Professor
Ph.D. Training Faculty
Department of Genetics
School of Medicine
Case Western Reserve University
Biomedical Research Building 619
2109 Adelbert Road
Cleveland, Ohio 44106-4955
Tel: (216) 368-6419
Fax: (216) 368-3432
E-mail: hua.lou@case.edu
http://cancer.cwru.edu
http://www.rnaresearch.org/


About Hua Lou

Hua Lou received a Ph.D. in Plant Molecular Biology at the University of Illinois at Urbaba-Champaign in 1993. After a postdoctoral fellowship at the University of Texas M.D. Anderson Cancer Center working with Dr. Robert Gagel, Dr. Lou moved to Baylor College of Medicine to work with Dr. Susan Berget as an Instructor, where she continued her work on alternative splicing. Dr. Lou joined the Department of Genetics in 2000.

Trainee achievements:
Hui Zhu (Ph.D., 2007): American Heart Association Predoctoral Fellowship (2004-2006); 2007 Harold M. Weintraub Graduate Student Award.
Hua-Lin Zhou (Ph.D.): American Heart Association Postdoctoral Fellowship (2007-2009); American Heart Association Third Year Postdoctoral Fellowship (2009-2010).
Victoria Barron (Ph.D. candidate): American Heart Association Predoctoral Fellowship (2008-2010).
Melissa Hinman (Ph.D. candidate): NINDS National Research Service Award, Predoctoral Fellowship (2010-2012).


Research

The research focus of my laboratory is to understand tissue-specific alternative splicing in mammals: the role of alternative splicing in development and the mechanisms that regulate tissue-specific alternative splicing. We are specifically interested in regulation of alternative splicing in neurons and heart tissues under both normal and pathological conditions. We combine genetic and biochemical approaches and use several model systems in our studies.

Role of alternative splicing in mammalian development

Alternative splicing plays an important role in the function of the gene product of a tumor suppressor gene, neurofibromatosis type 1 (NF1). Exon 23a of the NF1 pre-mRNA is an in-frame exon encoding 21 amino acids in the NF1 GTPase-activating protein (GAP) domain. This exon is alternatively included, producing two NF1 isoforms. The type I isoform does not contain this exon, while the type II isoform does. The ratio of the two isoforms varies in different tissues and during development. The type I isoform is predominantly expressed in neurons of the adult central nervous system and shows ten times higher activity in down-regulating Ras activity than the type II isoform.

To determine how changes of NF1 alternative splicing affects the role of NF1 in development, we are generating a mouse model in which the ratio of the two NF1 isoforms is drastically altered. By studying the phenotype of the knock-in mice, we will gain insight into regulated exon 23a expression in the development and function of the nervous system.

Mechanisms controlling alternative splicing of the NF1 exon 23a

Our recent studies demonstrate that alternative inclusion of exon 23a is controlled by at least three groups of RNA-binding proteins, Hu and CELF proteins as well as TIA-1/TIAR.

Hu proteins belong to a group of neuronal RNA-binding proteins that were originally cloned using antiserum from paraneoplastic neurologic disease (PND) patients and are antigens of the Hu syndrome triggered by small cell lung carcinomas. Hu proteins were shown to be required for neuronal differentiation in the mammalian nervous system. Our recent studies demonstrate that Hu proteins function as splicing regulators in addition to their well-established role in mRNA stability and translation. We have identified three splicing targets for Hu proteins including the NFI pre-mRNAs. We are currently studying the mechanisms that control the neuron-specific regulation of NF1 exon 23a skipping. We are particularly interested in how multiple splicing regulators coordinate with each other in this complex regulation. These studies will greatly increase our understanding of the mechanisms controlling alternative splicing in neurons. To study this regulation in its natural neuronal environment, we established a mouse embryonic stem cell system in which a specific lineage of neuronal cells can be differentiated from the ES cells. We also use mouse primary neurons for these studies.

Given the importance of the GAP function for NF1, it is conceivable that the splicing regulators modulating the expression of exon 23a can function as modifiers of the NF1 disease. See the following Department of Defense web site for more detailed description on this subject.
http://cdmrp.army.mil/highlights/default.htm#32

Regulation of co-transcriptional splicing

Recent studies indicate that for most eukaryotic genes, transcription and splicing are tightly coupled. The tight coupling of transcription and splicing predicts that transcription elongation rate can affect alternative splicing choice. In general, alternative exons are associated with suboptimal splicing signals and thus require more time than constitutive exons that are always included in the final mRNA to be recognized and defined. Studies in recent years have established that in general, increased transcription elongation rate promotes skipping of alternative exons while decreased elongation rate promotes inclusion.

Histone modification is associated with chromatin structure and speed of transcription elongation. Although several studies published during last year start to correlate histone modification and exon expression patterns, it is not clear how the two patterns are linked. We are currently investigating if and how splicing regulator proteins modulate transcription elongation rate, which in turn influence alternative splicing choice.

Splicing regulation by Fox-1/Fox-2 proteins

Previous research in my laboratory has identified Fox-1/Fox-2 as potent regulators promoting the neuronal splicing pathway of the calcitonin/calcitonin gene-related peptide (CGRP). Two functionally important binding sites of Fox-1/Fox-2 are located on either side of the 3' splice site of intron 3. We demonstrated that Fox proteins decrease binding of a basic splicing factor, U2AF to the polypyrimidine tract of the 3' splice site upstream from exon 4 to prevent formation of the earliest detectable spliceosomal complex E'.

Currently, we are investigating the protein composition of the E' pre-spliceosomal complex and determining the functional role of protein dimerization of Fox-1/Fox-2 in the regulation.


Selected Publications

Sharma A, Nguyen H, Geng C, Hinman MN, Luo G, Lou H (2014)
Calcium-mediated histone modifications regulate alternative splicing in cardiomyocytes.
Proc Natl Acad Sci U S A;111(46):E4920-8
See PubMed abstract

Hinman MN, Sharma A, Luo G, Lou H (2014)
Neurofibromatosis Type 1 Alternative Splicing is a Key Regulator of Ras Signaling in Neurons.
Mol Cell Biol;:
See PubMed abstract

Zhou HL, Luo G, Wise JA, Lou H (2013)
Regulation of alternative splicing by local histone modifications: potential roles for RNA-guided mechanisms.
Nucleic Acids Res;:
See PubMed abstract

Zhou HL, Geng C, Luo G, Lou H (2013)
The p97-UBXD8 complex destabilizes mRNA by promoting release of ubiquitinated HuR from mRNP.
Genes Dev;:
See PubMed abstract

Hinman MN, Zhou HL, Sharma A, Lou H (2013)
All three RNA recognition motifs and the hinge region of HuC play distinct roles in the regulation of alternative splicing.
Nucleic Acids Res;:
See PubMed abstract

Fleming VA, Geng C, Ladd AN, Lou H (2012)
Alternative splicing of the neurofibromatosis type 1 pre-mRNA is regulated by the muscleblind-like proteins and the CUG-BP and ELAV-like factors.
BMC Mol Biol;13:35
See PubMed abstract

Barron VA, Lou H (2012)
Alternative splicing of the neurofibromatosis type I pre-mRNA.
Biosci Rep;32(2):131-8
See PubMed abstract

Zhou HL, Hinman MN, Barron VA, Geng C, Zhou G, Luo G, Siegel RE, Lou H (2011)
Hu proteins regulate alternative splicing by inducing localized histone hyperacetylation in an RNA-dependent manner.
Proc Natl Acad Sci U S A;:
See PubMed abstract

Wang H., Molfenter J., Zhu H., Lou H. (2010)
Promotion of exon 6 inclusion in HuD pre-mRNA by Hu protein family members
Nucleic Acids Research;(In Press)
See PubMed abstract

Barron VA, Zhu H, Hinman MN, Ladd AN, Lou H (2009)
The neurofibromatosis type I pre-mRNA is a novel target of CELF protein-mediated splicing regulation.
Nucleic Acids Res;:
See PubMed abstract

Zhou HL, Lou H (2008)
Repression of prespliceosome complex formation at two distinct steps by Fox-1/Fox-2 proteins.
Mol Cell Biol;28(17):5507-16
See PubMed abstract

Hinman MN, Lou H (2008)
Diverse molecular functions of Hu proteins.
Cell Mol Life Sci;:
See PubMed abstract

Zhu H, Hinman MN, Hasman RA, Mehta P, Lou H (2007)
Regulation of Neuron-Specific Alternative Splicing of Neurofibromatosis Type 1 Pre-mRNA.
Mol Cell Biol;:
See PubMed abstract

Zhou HL, Baraniak AP, Lou H (2007)
Role for Fox-1/Fox-2 in mediating the neuronal pathway of calcitonin/calcitonin gene-related peptide alternative RNA processing.
Mol Cell Biol;27(3):830-41
See PubMed abstract

Zhu H, Zhou HL, Hasman RA, Lou H (2007)
Hu proteins regulate polyadenylation by blocking sites containing U-rich sequences.
J Biol Chem;282(4):2203-10
See PubMed abstract

Zhu H, Hasman RA, Barron VA, Luo G, Lou H (2006)
A nuclear function of Hu proteins as neuron-specific alternative RNA processing regulators.
Mol Biol Cell;17(12):5105-14
See PubMed abstract

Zhu H, Hasman RA, Young KM, Kedersha NL, Lou H (2003)
U1 snRNP-dependent function of TIAR in the regulation of alternative RNA processing of the human calcitonin/CGRP pre-mRNA.
Mol Cell Biol;23(17):5959-71
See PubMed abstract

Lou H, Gagel RF (2001)
Alternative ribonucleic acid processing in endocrine systems.
Endocr Rev;22(2):205-25
See PubMed abstract

Lou H, Helfman DM, Gagel RF, Berget SM (1999)
Polypyrimidine tract-binding protein positively regulates inclusion of an alternative 3'-terminal exon.
Mol Cell Biol;19(1):78-85
See PubMed abstract

Lou H, Gagel RF (1998)
Alternative RNA processing--its role in regulating expression of calcitonin/calcitonin gene-related peptide.
J Endocrinol;156(3):401-5
See PubMed abstract

Lou H, Gagel RF, Berget SM (1996)
An intron enhancer recognized by splicing factors activates polyadenylation.
Genes Dev;10(2):208-19
See PubMed abstract