Neural cell adhesion molecules in rat endocrine tissues and tumor cells: distribution and molecular analysis

The adhesive properties of neural cell adhesion molecules (NCAMs) can be modified by alternative splicing of the primary transcript or posttranslational modifications. In the present study, we describe distinct forms of alternative splicing and posttranslational modification of the extracellular domain of NCAM of various endocrine tissues and derived tumor cells of the rat. Using an antiserum detecting the immunoglobulin-like domains of NCAM as well as a monoclonal antibody recognizing the NCAM-specific polysialic acid (PSA), we observed a similar staining pattern in adrenals, pituitary, and neoplastic endocrine cells. In endocrine tumor cells [pheochromocytoma (PC12), insulinoma (RINA2), and pituitary tumor cells (GH3)], NCAM immunoreactivity was most intense at contact sites between the cells. The immunocytochemical data were substantiated by results of in situ hybridization histochemistry. Specifically, higher levels of NCAM mRNA were detected in the adrenal cortex than in the medulla. In the pituitary, NCAM mRNA was more abundant in the anterior and intermediate lobes than in the neural lobe. The sequence of NCAM mRNAs in endocrine cells was analyzed by polymerase chain reaction and S1 nuclease protection assays. We found that major exons 4-13 of the NCAM mRNA in endocrine tissues and related tumor cell lines were homologous to those in the brain. However, PC12, RINA2, and GH3 tumor cells; normal rat pituitaries; and adrenals contained different amounts of NCAM mRNA with an alternative extra exon, termed VASE (also called pi in mouse) between constitutive exons 7 and 8. In addition, in pituitaries, we detected an alternative exon in splice site a between the constitutive exons 12 and 13, termed a15, with or without an AAG triplett. These sites are thought to be important for the adhesive properties of NCAM. Therefore, these results suggest that modifications of NCAM may be important for adhesive interactions in normal and neoplastic endocrine cells

Leydig cells express neural cell adhesion molecules in vivo and in vitro

The neural cell adhesion molecule (NCAM) polypeptides are expressed by numerous tissues during embryonic development, where they are involved in cell-cell interactions. In the adult, NCAM expression is confined to a few cell types, including neurons and peptide-hormone-producing cells. Here we demonstrate that the Leydig cells of the adult rat, mouse, and hamster testes express NCAM as well. Western blotting showed that an NCAM of approximately 120 kDa was present in the adult testes of all three species investigated. This form was also found in freshly isolated mouse Leydig cells and in Leydig cells after 2 days in culture. After 4 days in culture, mouse Leydig cells expressed additional NCAM isoforms of approximately 140 and 180 kDa, indicating changes in alternative splicing of NCAM primary transcripts. Also, NCAM mRNA of all isoforms, as detected by S1-nuclease protection assays, increased with time in culture. The expression of the cell adhesion molecule NCAM by adult Leydig cells may explain the aggregation of Leydig cells in clusters in rodent testes, which could be a prerequisite for functional coordination of groups of Leydig cells. Furthermore, the presence of this neural and endocrine marker may indicate a closer relationship between Leydig cells and neural and peptide-hormone-producing cells than is considered to exist at the present time

NCAM-180, the large isoform of the neural cell adhesion molecule of the mouse, is encoded by an alternatively spliced transcript.

The three different isoforms (NCAM-180, -140 and -120) of the murine neural cell adhesion molecule are encoded by a single gene. The two smaller isoforms, NCAM-120 and -140 are generated by alternative RNA splicing of the primary transcript. We here report sequence data of a mouse cDNA clone reverse transcribed from NCAM mRNA containing an extra fragment of 801 nt. This cytoplasmic domain of NCAM-140 and codes for additional 267 amino acids. We conclude that the sequence presented represents the 3' portion of the 7.4 kb NCAM transcript, which is also generated by alternative splicing. Thus, this sequence probably encodes NCAM-180, a polypeptide with a Mr of 117,181

Chemical and Local Effects on Peptide Cysteine Reactivity

When a disulfide bond (R-S-S-R’) forms between the thiol groups of two cysteine amino acids this residue is known as cystine. Cysteine and cystine play a major role in protein structure. Disulfide bonds are usually formed in organisms by enzymes. The disulfide linkages can be scrambled in a protein or peptide with multiple disulfide bonds. This is associated with changes in protein structure and aggregation. Non-enzymatic disulfide formation presents challenges when studying cysteine containing peptides and proteins in the laboratory

Chemical and Local Effects on Peptide Cysteine Reactivity

When a disulfide bond (R-S-S-R’) forms between the thiol groups of two cysteine amino acids this residue is known as cystine. Cysteine and cystine play a major role in protein structure. Disulfide bonds are usually formed in organisms by enzymes. The disulfide linkages can be scrambled in a protein or peptide with multiple disulfide bonds. This is associated with changes in protein structure and aggregation. Non-enzymatic disulfide formation presents challenges when studying cysteine containing peptides and proteins in the laboratory

Nebenwirkungen und Verhalten von Noradrenalin und Adrenalin im Plasma beim intravenösen Thyroliberin-Test bei Personen mit normaler und gestörter Schilddrüsenfunktion

Peer Reviewe

Measures of Cysteine Acidity in Relation to Rates of Disulfide Formation

Cysteine and cystine residues play a major role in protein structure and the mediation of reactive oxygen species. Several neurodegenerative misfolding diseases (NMD) are associated with disulfide bond scrambling, including Alzheimer’s disease, Parkinson’s disease, prion-related disorders, and amyotrophic lateral sclerosis. Thioredoxin is a protein which is central to many pathways involving disulfides in biological systems. The difference in acidities of two active site cysteine residues of thioredoxin enables the reduction of disulfide bonds in proteins and peptides. The exact role the interactions between local residues and the functionality of the active site cysteines is unclear. We have found that disulfide bonds are formed between small cysteine containing peptides in solution in the absence of enzymes at different rates. Trends in the relative acidity of peptides were found to be congruent with trends in the rate of disulfide bond formation

Measures of Cysteine Acidity in Relation to Rates of Disulfide Formation

Cysteine and cystine residues play a major role in protein structure and the mediation of reactive oxygen species. Several neurodegenerative misfolding diseases (NMD) are associated with disulfide bond scrambling, including Alzheimer’s disease, Parkinson’s disease, prion-related disorders, and amyotrophic lateral sclerosis. Thioredoxin is a protein which is central to many pathways involving disulfides in biological systems. The difference in acidities of two active site cysteine residues of thioredoxin enables the reduction of disulfide bonds in proteins and peptides. The exact role the interactions between local residues and the functionality of the active site cysteines is unclear. We have found that disulfide bonds are formed between small cysteine containing peptides in solution in the absence of enzymes at different rates. Trends in the relative acidity of peptides were found to be congruent with trends in the rate of disulfide bond formation