Precerebellin-related genes and precerebellin 1 peptide in endocrine glands of the rat - pattern of their expression.

The hexadecapeptide cerebellin (CER) is derived from a larger protein, cerebellin 1 precursor protein (Cbln1). At present four precerebellins (Cbln1-4) are known. They are highly expressed in the brain, in particular in the cerebellum. Since CER is involved in regulating endocrine functions, present studies aimed to investigate, by means of molecular biology techniques (RT-PCR, QPCR, Western blotting) the expression of Cbln related genes and Cbln1 protein in classic endocrine glands of the rat. RT-PCR revealed the presence of Cbln1 and Cbln3 mRNAs in all endocrine glands tested; hypothalamus, anterior pituitary, thyroid, adrenal cortex, testis, ovary and pancreatic islets. Expression of Cbln2 gene was demonstrated only in the hypothalamus, anterior pituitary and adrenal cortex and in cerebral cortex, which was studied as a positive control organ. On the contrary, expression of Cbln4 gene was found only in the cerebral cortex. Using QPCR, the highest expression of Cbln1 gene was demonstrated in hypothalamus and pancreatic islets, a somewhat lower one in the anterior pituitary and thyroid, while the lowest was in adrenal cortex, testis and ovary. In general, the Cbln3 gene exhibited a similar pattern of expression, with the highest level in pancreatic islets and somewhat lower in the hypothalamus. Cbln2 gene expression was high in the hypothalamus, lower in the anterior pituitary and very low in adrenal cortex. In general, the pattern of Cbln1 protein expression was similar to that of Cbln1 mRNA. Further experiments aimed to check possible association of Cbln1 with cell membrane. Such association is suggested by differences in Cbln1 protein amount after extraction with RIPA and TRIS buffers. Bioinformatic methods predicting transmembrane topology (HMMTOP and SPLIT 4.0 servers) suggest transmembrane localisation of Cbln1, with transmembrane domain sequence responsible for the formation of an alpha-helix. These findings suggest possible physiological roles of Cbln related peptides not only in the cerebellum, but also in the endocrine system. However, their specific role as modulators of the endocrine system requires further investigations.