Expression and processing of the rooster protamine mRNA

In situ hybridization in immature and mature testis sections shows that the rooster protamine mRNA is transcribed in the post-meiotic stages of spermatogenesis. Two distinct populations of rooster protamine mRNA are expressed as determined by Northern blot analysis. Since there are two copies of the chicken protamine gene per haploid genome, the question was raised of whether the two mRNA populations correspond to the two different genes or was a result of differential mRNA processing. The fact that the two genes differ only in one nucleotide (one extra A in the polyadenylation signal in the second locus) and that random sequencing of several cDNA clones has revealed only one poly-A tail addition site favors the hypothesis that the differences are due to mRNA processing. This is supported by 3' S1 mapping which shows a single poly-A tail addition site, which in turn suggests that the heterogeneity is due to differences in the length of the poly-A tail. The latter is confirmed by RNAse H digestion of mRNA-Oligo-dT hybrids which shows that the two mRNA populations (470 +/- 20 nucleotide (nt) and 430 +/- 20 nt respectively) are converted to a single population of 345 +/- 15 nt in good accordance with the poly-A tail site determined by S1 mapping (347 nt). Thus one species has a poly-A tail of 145 nt appearing in round spermatids and the second, a shorter tail of 105 nt present at the later stages of elongated spermatids.