Pamela J. Mitchell Associate Professor of Biochemistry and Molecular Biology 125 Life Sciences Building, University Park, PA 16802 Phone: (814) 865-5802 Fax: (814) 863-7024 E-mail: pjm23@psu.edu B.A. in Biology, Rhode Island College M.A., M.S., and Ph.D. in Molecular Genetics, Columbia University Postdoctoral research at University of California, Berkeley Mitchell Lab Web Site

Developmental Gene Regulation in Drosophila and Mouse

Studies in my laboratory are directed towards understanding transcriptional regulation of gene expression during tissue morphogenesis. Much of our work has focused on the transcription factor AP-2 (aka AP-2α), a sequence-specific DNA binding protein that we have shown to be required for craniofacial, limb, body wall, and nervous system development in the mouse. In humans and mice, AP-2α belongs to a small family of transcription factors encoded by five genes. In mice, these are tfap2a, tfap2b, tfap2c, tfap2d, and tfap2e, encoding AP-2α, AP-2β, AP-2γ, AP-2δ, and AP-2ε, respectively).  In addition to known essential roles for AP-2α, AP-2β, and AP-2γ in embryonic development, AP-2 family transcription factors have also been implicated as tumor suppressors in breast and ovarian carcinomas and melanoma. To better understand the gene expression programs where AP-2 transcription factors function, we identified dAP-2, the sole homolog of AP-2 family genes in the fruitfly Drosophila melanogaster, and are using the versatile tools of Drosophila genetics to analyze its developmental regulation and functions.

dAP-2 exhibits the same DNA binding sequence specificity as mammalian AP-2 proteins, and its developmental expression pattern is an intriguing distillation of the expression patterns of murine AP-2 family members during mouse embryogenesis. Furthermore, an allelic series of 16 lethal dAP-2 point mutations generated in our laboratory confirms that major developmental roles of AP-2 have been conserved between flies and mice. dAP-2 null mutant die as newly eclosed/eclosing adults with a reduced proboscis, severely shortened legs, and defects in the protocerebral central complex, a brain region that controls locomotor behaviors. These defects are fundamentally analogous to the facial, limb, and nervous system defects we identified previously in AP-2a mutant mice and indicate that dAP-2 is an excellent model system for gaining insight into evolutionarily conserved regulatory pathways in limb and nervous system development.

Our research aims to 1) identify signaling pathways and transcription factors that direct cell type-specific expression of dAP-2 in leg and labial imaginal discs (primordia of the adult legs and proboscis) and developing central nervous system, 2) identify critical target genes regulated by dAP-2, and 3) test models identified in dAP-2 studies for relevance with regard to mammalian AP-2 family genes. As part of this work, we are using reporter gene analyses in transgenic flies together with in vitro molecular analyses to identify and characterize transcriptional enhancer-elements of the dAP-2 gene that are sufficient to direct specific aspects of the dAP-2 expression pattern in vivo. 

During leg development, dAP-2 is expressed at boundaries between leg segments (future joints) and is required cell-autonomously for joint formation and non-autonomously for growth of leg segments (Monge and Mitchell, 1998; Kerber et al, 2001; Monge et al, 2001). In recent studies, we have identified several independent enhancer elements in dAP-2 genomic regions sufficient to direct reporter gene expression to specific leg segment boundaries along the proximal-distal axis of the developing leg.  One of the enhancers directs expression to the presumptive tarsal joints and contains essential binding sites for Su(H), the transcription factor that, together with Notchintra, activates transcription of Notch target genes. In genetic mosaic experiments, we find that Su(H) is required for endogenous dAP-2 expression in presumptive tarsal joints, and is similarly required for expression of the tarsal joint-specific reporter transgene (Zou et al., in preparation). These data provide definitive evidence that dAP-2 is a direct target of Notch signaling in the developing leg.

Our studies of dAP-2 aim to advance understanding of AP-2 transcription factors and their roles during limb and nervous system development, and also shed light on how mis-regulation or mutation of these regulatory proteins affects tissue physiology and tumor progression.

Representative Publications:

• H. Schorle, P. Meier, M. Buchert, R. Jaenisch, and P.J. Mitchell. (1996). Transcription factor AP-2 essential for cranial closure and craniofacial development. Nature 381, 235-238.
• J. Richman and P.J. Mitchell. (1996). Craniofacial development: knock-out mice take one on the chin. Current Biology 6, 364-367.
• J.M. Fritschy, S. Brandner, A. Aguzzi, M. Koedood, B. Lüscher, and P.J. Mitchell. (1996). Brain cell type-specificity and gliosis-induced activation of the human cytomegalovirus immediate-early promoter in transgenic mice. J. Neurosci. 16, 2275-2282.
• I. Monge and P. J. Mitchell. (1998). DAP-2, the Drosophila homolog of transcription factor AP-2. Mech. Dev. 76:191-195.
• M. Maconochie, R. Krishnamurthy, S. Nonchev, P. Meier, M. Manzanares, P.J. Mitchell, R. Krumlauf. 1999. Regulation of Hoxa2 in cranial neural crest cells involves members of the AP-2 family. Development 126:1483-1494.
• Maytin E.V., Lin J.C., Krishnamurthy, R., Batchvarova N., Ron D., Mitchell, P.J. Habener J.F. (1999). Keratin 10 gene expression during differentiation of mouse epidermis requires transcription factors C/EBP and AP-2. Dev. Biol. 216:164.81.
• P.R. Kramer, G. Guerrero, R. Krishnamurthy, P.J. Mitchell, S. Wray. 2000. Ectopic expression of luteinizing hormone-releasing hormone and peripherin in the respiratory epithelium of mice lacking transcription factor AP-2alpha. Mech. Dev. 94:79-94.
• P.R. Kramer, R. Krishnamurthy R, P.J. Mitchell, S. Wray. 2000. Transcription factor activator protein-2 is required for continued luteinizing hormone-releasing hormone expression in the forebrain of developing mice. Endocrinology 141:1823-38.
• A. Antilla, J.K. Kellokoski, J.K., P.J. Mitchell, S. Saarikoski, K. Syrjanen, V. Kosma. 2000. Expression of transcription factor AP-2 predicts survival in epithelial ovarian cancer. Br. J. Cancer 82:1974-83.
• Monge I., Krishnamurthy R., Sims d., Hirth F., Spengler M., Kammermeier L., Reichert H., Mitchell, P.J. (2001). Drosophila transcription factor AP-2 in proboscis, leg and brain central complex development. Development 128:1239-52.
• Kerber B., Monge I., Mueller M., Mitchell, P.J., Cohen, S.M. (2001). The AP-2 transcription factor is required for joint formation and cell survival in Drosophila leg development. Development 128:1231-8.
• Stewart, H.J., A. Brennan, M. Rahman, M., G. Zoidl, P.J. Mitchell, K.R. Jessen, R. Mirsky. 2001. Developmental regulation and overexpression of the transcription factor AP-2, a potential regulator of the timing of Schwann cell generation. Eur. J. Neurosci.14:363-72.

P.J. Mitchell Abstracts

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