title: The expression of plasma membrane Ca(2+)-ATPase isoform 2 and its splice variants at sites A and C in the neonatal rat cochlea.
abstract: OBJECTIVE
  To study the expression of plasma membrane Ca(2+)-ATPase isoform 2 (PMCA2) and its alternative splicing at sites A (the first intracellular loop) and C (the C-terminal region) in the neonatal rat cochlea.
  
  
  METHODS
  The cochleae from rats postnatal day 3 to postnatal day 4 (P3-P4) were dissected, fixed, embedded, and sectioned. Meanwhile, the cochlear coils from neonatal rats were isolated and fixed. Using immunofluorescence staining, the expression of PMCA2 was respectively examined in the cochlear sections and cochlear coils. In addition, the total RNAs of basilar membrane (BM, including the organ of corti, the same below), spiral ganglion (SG), spiral ligament (SL, including SV, the same below), and the whole cochlea from neonatal rats were respectively extracted and reverse transcribed to cDNAs, then subjected to primers flanking site A or C in the PMCA2 gene using reverse transcription polymerase chain reaction (RT-PCR). Western blot was also applied to detect the expression of PMCA2 isoforms in the cochlear tissues.
  
  
  RESULTS
  We found that PMCA2 is strongly expressed in outer hair cell (OHC) bundles, SG, and stria vascularis (SV), weakly expressed in Reissner's membrane (RM), and occasionally expressed in inner hair cell (IHC) bundles. Moreover, w/a is the major splice form of PMCA2 present in hair cell bundles, z/b and z/c are the major splice forms of PMCA2 present in SG, and w/a and w/c are the major splice forms of PMCA2 present in SV. In the whole cochlea, variants w, y, and z were detected at site A, and variants a, b, and c were detected at site C. Using Western blot, variant a or b was also detectable in the same cochlear tissues mentioned above.
  
  
  CONCLUSIONS
  PMCA2 and its splice variants at sites A and C are differentially expressed in cochlear tissues of neonatal rat.
authors.affiliation: Department of Otolaryngology-Head and Neck Surgery, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, 1095 Jiefang Avenue, Wuhan, Hubei 430030, China.
authors.forename: Qingguo
authors.initials: Q
authors.lastname: Chen
authors.forename: Hanqi
authors.initials: H
authors.lastname: Chu
authors.forename: Xiaohui
authors.initials: X
authors.lastname: Wu
authors.forename: Yonghua
authors.initials: Y
authors.lastname: Cui
authors.forename: Jin
authors.initials: J
authors.lastname: Chen
authors.forename: Jianling
authors.initials: J
authors.lastname: Li
authors.forename: Liangqiang
authors.initials: L
authors.lastname: Zhou
authors.forename: Hao
authors.initials: H
authors.lastname: Xiong
authors.forename: Yan
authors.initials: Y
authors.lastname: Wang
authors.forename: Zhiyong
authors.initials: Z
authors.lastname: Li
mesh_terms: D017398:Alternative Splicing; D000818:Animals; D000831:Animals, Newborn; D015153:Blotting, Western; D003051:Cochlea / Q000378:metabolism / Q000502:physiology*; D018507:Gene Expression Regulation, Developmental; D006198:Hair Cells, Auditory / Q000378:metabolism / Q000502:physiology; D023421:Models, Animal; D053499:Plasma Membrane Calcium-Transporting ATPases / Q000235:genetics / Q000378:metabolism*; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012680:Sensitivity and Specificity
chemical_list: D020033:Protein Isoforms; D053499:Plasma Membrane Calcium-Transporting ATPases