Regulation of MBP function by its interaction with Ca2+-calmodulin and post-translational modifications

DURAND SANDRA E.; MAURICIO R. GALIANO; CHRISTOPHE BOSC; MARTA E. HALLAK

Huerta Grande, Cordoba

Congreso; I Reunión Conjunta de Sociedad Argentina de Investigación en Neurociencia (SAN) y Taller Argentino de Neurociencias (TAN); 2009

Sociedad Argentina de Investigación en Neurociencia (SAN) y Taller Argentino de Neurociencias (TAN)

Myelin basic protein (MBP) is essential for formation of myelin compact membranes in central nervous system. The interactions of MBP with different cell components are regulated by Ca2+-calmodulin and by various post-translational modifications. Since calmodulin (CaM) reverses the actin polymerisation and bundling effects of MBP, it might be a regulatory element, in concert with modifying enzymes such as kinases and deiminases, of the putative signaling role of MBP in vivo. In this regard, we study the MBP-CaM interaction by immobilized peptide arrays. There are six variants of MBP encoded by a single gene and generated by alternative splicing. The MBP isoform 1 contains peptide sequence encoded by all seven exons of the MBP gene representing the longest variant. For this reason, we performed the peptide array with this amino acid sequence and we find that CaM binds to three distinct domains that lay in exons involved in splicing and exon VII, common to all the isoforms. These CaM binding-sites differ in the strength of interaction. The strongest CaM binding-site is located in the C-terminal of 1, 2, 3 and 5 MBP isoforms. To analyze the relevance of these sites for the interaction of MBP with CaM we generated constructs that contain three, two, one or none CaM-binding site. Thus, we cloned the cDNA that codify to 1, 3, 4 and 6 MBP isoforms in a mammalian expression vector, plus their C-terminal deleted versions, to study their interaction with CaM in an oligodendroglial cell line that do not express MBP. In addition, each CaM binding-site has three serines and two arginines that can undergo the most common post-translational modifications of MBP such as phosphorylation or deimination respectively. Deimination of MBP has been correlated with early development, demyelination process and disease severity in multiple sclerosis (MS). Further study of the regulation of MBP by Ca2+-calmodulin and post-translational modifications will help to understand its role in signaling for oligodendrocyte differentiation and myelin formation, and its involvement in the pathogenesis of MS.