Isolation and analysis of the 3 '-untranslated regions of the human relaxin H1 and H2 genes

The human has two relaxins, termed H1 and H2, both of which are biologicall y active and coexpressed in the decidua, placenta and prostate; in the corp us luteum, the main source of circulating relaxin, only the H2 form is expr essed. The reasons for this differential expression of the relaxin genes ar e unknown. The possibility that their 3'-untranslated regions (UTRs) contri bute to this differential expression by affecting their mRNA stabilities wa s investigated. Thus the 3'-UTRs of both relaxin genes were isolated throug h a combined 3'-rapid amplification of cDNA ends-PCR (RACE-PCR) using poly (A)(+) RNA from human decidua, placenta, prostate and corpus luteum. The se quences obtained for each 3'-UTR were identical in the tissues examined, we re AT-rich (72%) and showed 91% homology between relaxin H1 and H2 when max imally aligned to include several gaps, the significance of which is unknow n. Relaxin H1 has two, and relaxin H2 has one, poly (A)(+) signal, in addit ion to one cytoplasmic polyadenylation element 30 nucleotides upstream of t his. The mRNA levels of relaxin H1 and H2 in the prostate adenocarcinoma LN CaP.FGC cell line were determined by quantitative competitive RT-PCR. Relax in H1 had a 10-fold greater number of molecules (similar to 2.5 x 10(7)) pe r mu g of total RNA than relaxin H2 (similar to 2.5 x 10(6)). The stability of relaxin H1 and H2 mRNAs were compared in LnTCaP cells treated with the transcription inhibitor actinomycin D (10 mM) for 0, 1, 2, 4, 8, 10, 14, or 24 h. Half-lives of 3.17 days for relaxin H1 mRNA and 11.4 h for relaxin H 2 mRNA were obtained from semi-logarithmic plots. Thus both mRNAs are relat ively stable; however, relaxin H1 mRNA is considerably more stable than rel axin H2, at least in LNCaP cells. This difference in their mRNA stability m ay partly explain the greater level of expression of relaxin H1 in these ce lls.