Show simple item record

dc.contributor.authorGeist Hauserman, Janelle
dc.date.accessioned2019-09-18T15:45:13Z
dc.date.available2019-09-18T15:45:13Z
dc.date.issued2019
dc.identifier.urihttp://hdl.handle.net/10713/10903
dc.description2019en_US
dc.descriptionBiochemistry and Molecular Biologyen_US
dc.descriptionUniversity of Maryland, Baltimoreen_US
dc.descriptionPh.D.en_US
dc.description.abstractMyosin Binding Protein-C (MyBP-C) comprises a family of proteins with structural and regulatory roles in muscle. There are three MyBP-C isoforms in the family, encoded by different genes. Although the isoforms share significant structural and sequence homology, slow skeletal MyBP-C (sMyBP-C), encoded by MYBPC1, is unique as it is heavily spliced in both the NH2 and COOH-termini. To study the role of sMyBP-C in healthy, adult skeletal muscles, in vivo gene transfer and CRISPR plasmids were used to knock down sMyBP-C. Decreased sMyBP-C levels resulted in significantly decreased levels of thick, but not thin, filament proteins. The reduced levels of thick filament proteins were accompanied by disorganized A- and M-bands. Moreover, examination of the contractile activity of treated muscles demonstrated that downregulation of sMyBP-C resulted in significantly decreased force production and velocities of contraction and relaxation. In addition to the extensive exon shuffling that takes place in the NH2-terminus of sMyBP-C, it also undergoes PKA and PKC mediated phosphorylation within two motifs, which flank the first Ig domain of the protein. Recombinant NH2-terminal sMyBP-C phosphomimetic peptides were tested in co-sedimentation and in vitro motility assays, indicating that phosphorylation of sMyBP-C variants regulates actomyosin binding and sliding velocity. Mutations in MYBPC1 have been implicated in the development of distal arthrogryposis, while four recently discovered mutations (Y247H, E248K, L259P, and L263R) co-segregate with the development of a new myopathy characterized by muscle weakness, hypotonia, skeletal deformities, and tremor. In vitro studies and computational modeling suggest altered myosin binding and/or protein instability for the four mutations. Further in vivo evaluation of the E248K mutation in a heterozygous knock-in mouse model revealed significant biochemical, morphological, and behavioral deficits compared to wild type littermates. Additionally, functional assessment of heterozygous E248K muscles demonstrated decreased force and power production, as well as decreased cross bridge cycling kinetics, indicating the tremor may begin at the level of the sarcomere. My studies therefore reveal that sMyBP-C has important structural and regulatory roles within the sarcomere, is modulated through phosphorylation, and that novel MYBPC1 mutations lead to the development of myopathy and tremor that is of myogenic origin.en_US
dc.language.isoen_USen_US
dc.subjectCRISPRen_US
dc.subjectmyosin binding protein-Cen_US
dc.subject.meshGene Knockdown Techniquesen_US
dc.subject.meshMuscle, Skeletalen_US
dc.subject.meshTremoren_US
dc.titleFunctional characterization of Myosin Binding Protein-C slow in health and diseaseen_US
dc.typedissertationen_US
dc.date.updated2019-09-12T22:04:30Z
dc.language.rfc3066en
dc.contributor.advisorKontrogianni-Konstantopoulos, Aikaterini
dc.description.embargo03/12/2020
refterms.dateFOA2019-09-18T15:45:14Z


Files in this item

Thumbnail
Name:
GeistHauserman_umaryland_0373D ...
Size:
10.02Mb
Format:
PDF

This item appears in the following Collection(s)

Show simple item record