Plexin B1 is repressed by oncogenic B-Raf signaling and functions as a tumor suppressor in melanoma cells.

Human melanomas show oncogenic B-Raf mutations, which activate the B-Raf/MKK/ERK cascade. We screened microarrays to identify cellular targets of this pathway, and found that genes upregulated by B-Raf/MKK/ERK showed highest association with cell-cycle regulators, whereas genes downregulated were most highly associated with axon guidance genes, including plexin-semaphorin family members. Plexin B1 was strongly inhibited by mitogen-activated protein kinase signaling in melanoma cells and melanocytes. In primary melanoma cells, plexin B1 blocked tumorigenesis as measured by growth of colonies in soft agar, spheroids in extracellular matrix and xenograft tumors. Tumor suppression depended on residues in the C-terminal domain of plexin B1, which mediate receptor GTPase activating protein activity, and also correlated with AKT inhibition. Interestingly, the inhibitory response to plexin B1 was reduced or absent in cells from a matched metastatic tumor, suggesting that changes occur in metastatic cells which bypass the tumor-suppressor mechanisms. Plexin B1 also inhibited cell migration, but this was seen in metastatic cells and not in matched primary cells. Thus, plexin B1 has tumor-suppressor function in early-stage cells, although suppressing migration in late-stage cells. Our findings suggest that B-Raf/MKK/ERK provides a permissive environment for melanoma genesis by modulating plexin B1.

Selective down-regulation of progesterone receptor isoform B in poorly differentiated human endometrial cancer cells: implications for unopposed estrogen action.

The uterine endometrium responds to unopposed estrogen stimulation with rapid cell proliferation. Progesterone protects the endometrium against the hyperplastic effects of estradiol (E2) through progesterone receptors (PRs), of which two isoforms are expressed: human (h) PRA and PRB. hPRB has a longer NH2 terminus and may function differently from hPRA. Thus, the relative expression of hPRA:hPRB is likely to be important for the action of progesterone. We hypothesized that the hPRA:hPRB ratios may be abnormal in endometrial cancer, leading to a lack of normal progesterone protection against the growth-promoting effects of E2. To test this hypothesis, well-differentiated Ishikawa endometrial cancer cells were compared to poorly differentiated Hec50 and KLE cells. Reverse transcription-PCR was chosen as a sensitive method to detect transcripts for the two forms of PR. The relative expression of PR isoforms under hormonal stimulation was determined by Western blotting. Transient transfections of hPRA and hPRB into endometrial cells allowed the evaluation of the transcriptional activity of each isoform independently on reporter gene transcription under the control of a simple progesterone response element-containing promoter. The effect of coexpressing the estrogen receptor on PR expression was also studied. Ishikawa cells (well-differentiated) express both hPRA and hPRB. Both isoforms, but predominantly hPRB, are up-regulated by E2 and not by tamoxifen or the pure antiestrogen ICI 182,780. Hec50 and KLE cells (poorly differentiated) express only hPRA. No hPRB is present in the poorly differentiated cells, and it is not induced by estrogen receptor expression and/or estrogen treatment. In all cells, hPRB expression, whether endogenous or produced as a result of transfection, acts as a stronger transcription factor than hPRA on a simple progesterone-dependent promoter. We speculate that down-regulation of hPRB may predict for poorly differentiated endometrial cancers that do not respond to progestin therapy.