The tissue-specific expession of the two murine alpha-amylase loci Amy-I
+
PPorotid AMV-1 a
1--- 1--- 1---1--- IL __
PPoncreos AMV-20
~~-1==---~_:~=== +
Fig. 7.11 Transcription termination in mouse Amy-I and Amy-2 genes. Major and minor poly A sites are indicated by large and small arrows, respectively. Most Amy- I transcripts are terminated close to the poly A sites. A few transcripts extend as much as 3 kb downstream of this si te. Transcription termination is identical in parotid and liver, suggesting that it does not depend on the choice of the promoter. Virtually all Amy-2 transcripts extend at least 2 kb downstream of Lhe major poly A site and are terminated at multiple positions between 2 kb and 4 kb downstream of this site.
and Amy-2 is summarized in Fig. 7.12. Our results concerning the structure and expression of alpha-amylase genes document that they are part of a very dynamic gene family. During evolution, multiple gene copies have arisen through duplication and diversification of a common ancestral gene, prob-ably active only in the pancreas of vertebrates (Meisler and Gumucio 1985b).
Some of the copies may have acquired different tissue specificities by several mechanisms, including point mutations, promoter insertions via trans-posable elements and/or promoter duplication, and diversification of the duplicated control element. Other gene copies have suffered modifications that inactivated them. The multiple pseudogenes observed in several mouse strains (see above) as well as in rats (MacDonald et al. 1980) are remnants of such genes.
Modulation of alpha-amylase expression is regulated mainly by two mechanisms in the three tissues that produce this enzyme: differential strength of tissue-specific promoters and, in the pancreas, which is the most active alpha-amylase producer, the utilization of multiple gene copies (gene dosage). Since transcription rates can largely account for the differential accumulation of alpha-amylase mRNAs in the three expressing tissues, we consider it unlikely that these transcripts vary significantly in stability.
Transcription from the two Amy-I promoters is not only tissue specific but also stage specific during parotid gland differentiation. All the expression studies reported in this paper are consistent with the speculation that the weak promoter, PL, is much less stringently controlled than the
parotid-'
'Fig. 7.12 Summary of transcription ofmurine Amy-1 and Amy-2 genes. The Amy-1 and Amy-2 transcripts observed in the different alpha-amylase-expressing tissues are shown by arrows. The varying thickness of the arrows reflects differenL promoter strengths. All four Amy-2 aJ1eles of mouse strain A/J are assumed to be transcrip-tional'ly active although tbjs has only be rigorously shown for two gene copies (see text).
specific strong promoter Pp. Actually, it appears quite likely that the activity of the former promoter merely reflects a transcriptionally permissive chro-matin structure of Amy- I. In this context it is interesting to note that Amy- I is in an accessible chromatin domain in the pancreas. Is the converse true for Amy-2 in parotid and/or liver? Recent chromatin studies aimed at answering this question have shown that, in these two tissues, Amy-2 chromatin is equally as resistant to digestion with DNAase I as it is in spleen, which does not express either of the two alpha-amylase loci (Schibler and Pittet, unpub-lished observation). One may argue, therefore, that sequences located within the Amy-I locus are required for opening the chromatin of both loci."1f cor-rect, this hypothesis would imply that during differentiation of all alpha-amylase-expressing tissues, the alpha-amylase chromatin starts opening at identical sequences but closes downstream of Amy- I in parotid and liver, and downstream of the Amy-2 complex in the pancreas. According to this model, the activation of Amy-2 during pancreatic development may require far upstream Amy-I sequences. It will be very exciting to test this hypothesis in transgenic mice.
Acknowledgements
We are grateful to A.-C. Pittet and R. Bovey for excellent technical assist-ance. P. H. S. acknowledges an NIH postdoctoral fellowship and M. C. a predoctoral fellowship from the Brazilian Research Council CNPq.
We thank K. Gorski for reading the manuscript. These studies were sup-ported by grants from the Swiss National Science foundation to U. S., P. KW. and 0 . H.
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