title: Differential expression of utrophin-A and -B promoters in the central nervous system (CNS) of normal and dystrophic mdx mice.
abstract: Utrophin (Utrn) is the autosomal homolog of dystrophin, the Duchene Muscular Dystrophy (DMD) locus product and of therapeutic interest, as its overexpression can compensate dystrophin's absence. Utrn is transcribed by Utrn-A and -B promoters with mRNAs differing at their 5' ends. However, previous central nervous system (CNS) studies used C-terminal antibodies recognizing both isoforms. As this distinction may impact upregulation strategies, we generated Utrn-A and -B promoter-specific antibodies, Taqman Polymerase chain reaction (PCR)-based absolute copy number assays, and luciferase-reporter constructs to study CNS of normal and dystrophic mdx mice. Differential expression of Utrn-A and -B was noted in microdissected and capillary-enriched fractions. At the protein level, Utrn-B was predominantly expressed in vasculature and ependymal lining, whereas Utrn-A was expressed in neurons, astrocytes, choroid plexus and pia mater. mRNA quantification demonstrated matching patterns of differential expression; however, transcription-translation mismatch was noted for Utrn-B in caudal brain regions. Utrn-A and Utrn-B proteins were significantly upregulated in olfactory bulb and cerebellum of mdx brain. Differential promoter activity, mRNA and protein expressions were studied in cultured C2C12, bEnd3, neurons and astrocytes. Promoter activity ranking for Utrn-A and -B was neurons > astrocytes > C2C12 > bEnd3 and bEnd3 > astrocytes > neurons > C2C12, respectively. Our results identify promoter usage patterns for therapeutic targeting and define promoter-specific differential distribution of Utrn isoforms in normal and dystrophic CNS.
authors.affiliation: Department of Physiology and Pennsylvania Muscle Institute, University of Pennsylvania School of Medicine, A-601 Richards Building, 3700 Hamilton Walk, Philadelphia, Pennsylvania 19104-6085, USA.
authors.forename: Santhosh M
authors.initials: SM
authors.lastname: Baby
authors.forename: Sasha
authors.initials: S
authors.lastname: Bogdanovich
authors.forename: Gabriel
authors.initials: G
authors.lastname: Willmann
authors.forename: Utpal
authors.initials: U
authors.lastname: Basu
authors.forename: Olga
authors.initials: O
authors.lastname: Lozynska
authors.forename: Tejvir S
authors.initials: TS
authors.lastname: Khurana
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001921:Brain / Q000098:blood supply / Q000378:metabolism*; D002460:Cell Line; D002478:Cells, Cultured; D003964:Diaphragm / Q000378:metabolism; D016189:Dystrophin / Q000172:deficiency / Q000378:metabolism; D008168:Lung / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D018101:Mice, Inbred mdx; D018482:Muscle, Skeletal / Q000378:metabolism; D009136:Muscular Dystrophies / Q000235:genetics / Q000378:metabolism*; D009206:Myocardium / Q000378:metabolism; D011401:Promoter Regions, Genetic; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D013116:Spinal Cord / Q000378:metabolism*; D049411:Utrophin / Q000235:genetics / Q000378:metabolism*
chemical_list: D016189:Dystrophin; D020033:Protein Isoforms; D012333:RNA, Messenger; C483600:Utrn protein, mouse; D049411:Utrophin; C551344:utrophin B, mouse