Foxo3a regulation of GALT expression provides a possible mechanism for the ovarian defect in mice expressing only the short form of the PRL receptor

HALPERIN J, BOWEN-SHAUVER J, MORAN T, LESLIE N, BINART N, KELLY P AND GIBORI G.

Chicago, USA

Jornada; NIH Annual Specialized Cooperative Centers Program in Reproductive Research (SCCPRR) Meeting; 2005

National Institutes of Health (NIH)

<!-- /* Font Definitions */ @font-face {font-family:"Cambria Math"; panose-1:2 4 5 3 5 4 6 3 2 4; mso-font-charset:0; mso-generic-font-family:roman; mso-font-pitch:variable; mso-font-signature:-1610611985 1107304683 0 0 159 0;} @font-face {font-family:Calibri; panose-1:2 15 5 2 2 2 4 3 2 4; mso-font-charset:0; mso-generic-font-family:swiss; mso-font-pitch:variable; mso-font-signature:-1610611985 1073750139 0 0 159 0;} /* Style Definitions */ p.MsoNormal, li.MsoNormal, div.MsoNormal {mso-style-unhide:no; mso-style-qformat:yes; mso-style-parent:""; margin:0in; margin-bottom:.0001pt; mso-pagination:widow-orphan; font-size:12.0pt; font-family:"Times New Roman","serif"; mso-fareast-font-family:"Times New Roman";} .MsoChpDefault {mso-style-type:export-only; mso-default-props:yes; font-size:10.0pt; mso-ansi-font-size:10.0pt; mso-bidi-font-size:10.0pt; mso-ascii-font-family:Calibri; mso-fareast-font-family:Calibri; mso-hansi-font-family:Calibri;} @page Section1 {size:8.5in 11.0in; margin:1.0in 1.0in 1.0in 1.0in; mso-header-margin:.5in; mso-footer-margin:.5in; mso-paper-source:0;} div.Section1 {page:Section1;} --> Alternative splicing of the prolactin receptor (PRLR) generates two isoforms, the short (RS) and long (RL) form.  The mechanism by which PRL signals through the RL has been extensively investigated.  However, little is known about PRL signaling through the RS.  To ascertain the role of the RS in the ovary, PRLR null mice expressing a transgenic RS construct were generated.  A severe ovarian defect and premature ovarian failure were found in these mice.  The ovarian phenotype in the RS transgenic mice is identical to phenotypes found in both Foxo3a null mice and in women carrying mutations of galactose-1-phosphate uridyl transferase (GALT).  Foxo proteins are forkhead transcription factors known to regulate genes involved in glucose metabolism while GALT is an enzyme that converts galactose-1-P to glucose.  Deficiencies in GALT lead to galactosemia and premature ovarian failure.  To investigate whether Foxo3a expression is regulated by PRL signaling through the RS, ovaries from PRLR null and RS transgenic mice were subjected to microarray analysis, RT-PCR, or Real Time PCR.  We found that expression of RS in the ovary leads to significant down-regulation of Foxo3a expression.  HepG2 cells were co-transfected with the full-length 3.3kb GALT promoter along with either a constitutively active (CA) Foxo3a construct or an empty vector in order to determine whether Foxo3a regulates GALT expression. Transfection studies showed a marked up-regulation of the GALT promoter by Foxo3a.  A 5’ deletion analysis of the GALT promoter was performed to distinguish the Foxo3a regulatory sites from the six putative Foxo3a binding sites.  Co-transfection with the truncated promoter-reporter constructs and CA- Foxo3a revealed a response element, which significantly up-regulated GALT gene expression, at –1011 bp upstream of the ATG site.  In summary, the results indicate that PRL, acting through the short form of the receptor, down-regulates the expression of Foxo3a affecting the transcription of GALT.  Foxo3a-GALT interaction provides a possible mechanism for the severe follicular impairment in females expressing RS.  Supported by NIH HD11119 and U54 HD 40093.