The role of MAP kinases in the regulation of steroid hormone synthesis will be studied in a insect model system, the prothoracic gland of the tobacco hornworm Manduca sexta. Episodic increases in ecdysteroid synthesis by the gland coordinate gene expression during development. Ecdysteroid synthesis is acutely regulated by a brain neuropeptide hormone, prothoracicotropic hormone (PTTH), in a system with considerable homology to vertebrate systems such as the ACT'H-adrenal cortex axis. These studies will expand upon previous work on signal transduction and steroidogenesis, with the ultimate goal of understanding the extrinsic and intrinsic controls of ecdysteroid synthesis. Preliminary experiments revealed that FTFH evokes a rapid increase in the phosphorylation of the MAP kinases known as ERKs (extracellular signal-regulated kinases). Inhibitors of ERK phosphorylation caused at least a partial inhibition of PTTH-stimulated ecdysteroid synthesis. Biochemical, physiological, pharmacological and molecular techniques will be used to rigorously determine the degree to which PTTH-dependent ecdysteroid and protein synthesis are dependent on activated ERKs. A second goal of the project is to determine as far as possible the upstream effectors of ERK activation and the downstream targets of ERKs. Among the candidate ERK targets, modulators of general and specific protein synthesis are of special interest because PTTH-stimulated ecdysteroid synthesis requires de novo translation. Upstream of ERKs, determining the possible involvement of cyclic AMP-dependent protein kinase (PKA) is of prime concern, since PKA activity is required for maximal PTTH-stimulated ecdysteroid synthesis but new data suggest that tyrosine kinases may also be involved. Feedback of ecdysteroids on the prothoracic gland down-regulates the ability of the gland to further synthesize ecdysteroids. The third goal of the project will address the possible ecdysteroid regulation of ERK levels as one way by which this negative feedback is accomplished. These studies will shed light on how intracellular regulatory pathways interact and should be of special interest to the endocrine field because prothoracic glands are not proliferative. They thus provide a system in which the acute stimulation of steroid synthesis is uncoupled from cell division.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
1R01GM063198-01
Application #
6322403
Study Section
Special Emphasis Panel (ZRG1-TMP (01))
Program Officer
Cole, Alison E
Project Start
2001-04-01
Project End
2005-03-31
Budget Start
2001-04-01
Budget End
2002-03-31
Support Year
1
Fiscal Year
2001
Total Cost
$180,992
Indirect Cost
Name
University of North Carolina Chapel Hill
Department
Biology
Type
Schools of Arts and Sciences
DUNS #
078861598
City
Chapel Hill
State
NC
Country
United States
Zip Code
27599
Rybczynski, Robert; Gilbert, Lawrence I (2006) Protein kinase C modulates ecdysteroidogenesis in the prothoracic gland of the tobacco hornworm, Manduca sexta. Mol Cell Endocrinol 251:78-87
Ono, Hajime; Rewitz, Kim F; Shinoda, Tetsuro et al. (2006) Spook and Spookier code for stage-specific components of the ecdysone biosynthetic pathway in Diptera. Dev Biol 298:555-70
Rewitz, Kim F; Rybczynski, Robert; Warren, James T et al. (2006) Identification, characterization and developmental expression of Halloween genes encoding P450 enzymes mediating ecdysone biosynthesis in the tobacco hornworm, Manduca sexta. Insect Biochem Mol Biol 36:188-99
Rewitz, Kim F; Rybczynski, Robert; Warren, James T et al. (2006) Developmental expression of Manduca shade, the P450 mediating the final step in molting hormone synthesis. Mol Cell Endocrinol 247:166-74
Rewitz, K F; Rybczynski, R; Warren, J T et al. (2006) The Halloween genes code for cytochrome P450 enzymes mediating synthesis of the insect moulting hormone. Biochem Soc Trans 34:1256-60
Fellner, Susan K; Rybczynski, Robert; Gilbert, Lawrence I (2005) Ca2+ signaling in prothoracicotropic hormone-stimulated prothoracic gland cells of Manduca sexta: evidence for mobilization and entry mechanisms. Insect Biochem Mol Biol 35:263-75
Rybczynski, R; Gilbert, L I (2003) Prothoracicotropic hormone stimulated extracellular signal-regulated kinase (ERK) activity: the changing roles of Ca(2+)- and cAMP-dependent mechanisms in the insect prothoracic glands during metamorphosis. Mol Cell Endocrinol 205:159-68
Rybczynski, R; Bell, S C; Gilbert, L I (2001) Activation of an extracellular signal-regulated kinase (ERK) by the insect prothoracicotropic hormone. Mol Cell Endocrinol 184:1-11