(1) Characterization of cDNA clones selected from a library of Xenopus anterior pituitary cDNAs by the mouse POMC cDNA is continuing. The clone containing the REM1 repetitive sequence is being sequenced by use of internal primers. Another clone may contain regulatory sequences for Xenopus POMC and is expected also to contain some POMC coding sequence. (2) In the neuroblastoma project, several cDNA clones with quantitative or qualitative specificity for either differentiated or undifferentiated cells are being characterized by Northern, Southern, and dot blots, nuclease protection and primer extension experiments, and sequencing. Interesting inserts are being transferred to a riboprobe-like vector to facilitate these studies, and a genomic library from differentiated NS20Y neuroblastoma cells has been prepared. (3) The cloned cDNA GammaChAT7, presumed to correspond to a portion of human choline acetyltransferase has been sequenced using internal oligonucleotide primers. Efforts to determine definitively whether this cDNA represents ChAT are continuing while, in the meantime, additional clones are being sought by screening with synethetic oligonucleotide probes whose sequences have been deduced from two cyanogen bromide peptides sequenced from the human placental enzyme. (4) A new project has been initiated in which cDNA clones are being obtained which are unique to, or enriched in, the subchronic reaction around a stab wound in rat cerebral cortex. Of interest are all lesion-induced mRNAs, but particularly that for a trophic factor expressed preferentially in lesioned versus control brain, which can be detected by its capacity to promote the survival of cultured sympathetic neurons. The initial characterization of a sampling of the library showed that the inserts, although small, correspond to brain mRNAs. Characterization of these clones, as well as a search for the trophic factor(s) and preparation of another cDNA library, are in progress. (5) The cDNA coding sequence for mouse POMC was subcloned into PUC19 expression vector, and several clones which express large amounts of this protein have been obtained.
