The transactivation‐competent carboxyl‐terminal domain of AF‐9 is expressed within a sexually dimorphic transcript in rat pituitary

Abstract

We have investigated the biological role of the cellular counterpart of the leukemogenic AF-9 gene by cloning the rat AF-9 (rAF-9) cDNA and defining the regulation of an anterior pituitary-specific rAF-9 transcript that is expressed in a sexually dimorphic manner. Expression of this transcript is down-regulated after puberty in females and can be subsequently up-regulated in adults by ovariectomy. Hormone replacement studies have provided direct evidence that rAF-9 mRNA expression is suppressed by estrogen. Mapping the 1.9 kb anterior pituitary transcript has shown that it corresponds in size to the rAF-9 cDNA clone, which contains an open reading frame (ORF) that is truncated compared with the human AF-9 ORF, but encodes a previously defined transcriptional activation domain. Thus, the cellular AF-9 gene is alternatively expressed in a manner that reflects the presence of translocated, functionally active (oncogenic) AF-9 sequences in leukemias. Using a novel antisera raised against a rAF-9 peptide, we have also demonstrated tissue- and sex-specific expression of a nuclear 41 kDa anterior pituitary protein and have localized this protein to a major population of growth hormone synthesizing cells. By localizing the expression and defining the physiological regulation of rAF-9, our studies have provided novel insights into the AF-9 gene that will facilitate an understanding of both oncogenic and endocrine roles.

The polycomb protein MPc3 interacts with AF9, an MLL fusion partner in t(9;11)(p22;q23) acute leukemias

“…Particularly high levels of expression are noted in the spleen, and expression during embryogenesis appears highest in early embryos (Figure 4). These results are in general agreement with analyses of AF9 transcripts in humans and rats by Northern blot (Iida et al, 1993;Morgan et al, 2000).…”

Oestrogenic regulation of an egr-1 transgene in rat anterior pituitary

“…Within the anterior lobe, transgene transcripts are further restricted to a sub-population of cells indicating that the transgene is also able to mediate cell-type-specific transcriptional control. Thus, using a previously validated (Morgan et al 2000) dual ISH/ immunohistochemical technique we have demonstrated co-localization of the transgene transcript with LH -subunit immunoreactivity. This finding reflects our demonstration of Egr-1 protein in rat gonadotrophs (Knight et al 2000), and indicates a remarkable fidelity of cell-type-specific expression that is conferred by egr-1 sequences in the transgene.…”

“…1D). Dual ISH/immunocytochemistry (Allen et al 1997, Morgan et al 2000 using the LH -subunit antiserum in combination with the antisense GFP probe revealed co-localization of LH -subunit immunoreactivity with transgene mRNA (Fig. 1D).…”

“…Dual ISH/immunocytochemistry was performed using a previously described protocol (Allen et al 1997, Morgan et al 2000 adapted for gonadotrophs. All solutions used in the initial immunocytochemical step were prepared in diethylpyrocarbonate-treated water, and also contained 0·1 U/µl RNasin ribonuclease inhibitor (Promega).…”

cDNA Array Analysis of Pineal Gene Expression Reveals Circadian Rhythmicity of the Dominant Negative Helix‐Loop‐Helix Protein‐Encoding Gene, Id‐1

“…Atlas Rat 1.2 cDNA Expression Arrays (Clontech Laboratories, Palo Alto, CA, USA), consisting of rat cDNA fragments from 1176 genes, were used in this study. Total RNA was extracted from a pool of 2 rat pineals sampled at either 12.00 h (L) or 24.00 h (D) using a standard protocol (16), and DNAse I‐treated according to the Atlas protocol. RNA quality was then assessed by agarose gel electrophoresis.…”