Translation and assembly of CABYR coding region B in fibrous sheath and restriction of calcium binding to coding region A.
CABYR is a highly polymorphic, sperm flagellar calcium-binding protein that is tyrosine as well as serine/threonine phosphorylated during capacitation. Six alternative splice variants of human CABYR (I-VI) have previously been identified, involving two coding regions, CR-A and CR-B, separated by an intervening stop codon. It is presently unknown if proteins encoded by the predicted coding region B of CABYR are translated during spermiogenesis, where they localize, or which CABYR isoforms bind calcium. Immunofluorescent and electron microscopic studies using polyclonal antibodies generated to the recombinant c-terminal 198 aa CABYR-B localized the isoforms containing CABYR-B to the ribs and longitudinal columns of the fibrous sheath in the principal piece of the flagellum. Antisera to recombinant CABYR-A and CABYR-B proteins recognized distinct populations of CABYR isoforms encoded by either CR-A alone and/or CR-B as well as a common population of CABYR isoforms. Only the recombinant CABYR-A and not the CABYR-B bound calcium in vitro, which is consistent with the hypothesis that CABYR-A is the only form that binds calcium in sperm. These observations confirmed that, despite the presence of the stop codon in CR-A, splice variants containing CR-B are expressed during spermiogenesis and assemble into the fibrous sheath of the principal piece; however, calcium binding occurs only to those CABYR isoforms containing CABYR-A.
Kim
Young-Hwan
YH
Center for Research in Contraceptive and Reproductive Health, Department of Cell Biology, University of Virginia, P.O. Box 800732, Charlottesville, VA 22908, USA.
Jha
Kula N
KN
Mandal
Arabinda
A
Vanage
Geeta
G
Farris
Erin
E
Snow
Phillip L
PL
Klotz
Ken
K
Naaby-Hansen
Soren
S
Flickinger
Charles J
CJ
Herr
John C
JC
D017398:Alternative Splicing; D000818:Animals; D000906:Antibodies; D001483:Base Sequence; D002118:Calcium / Q000378:metabolism; D002135:Calcium-Binding Proteins / Q000737:chemistry / Q000235:genetics* / Q000276:immunology / Q000378:metabolism*; D003001:Cloning, Molecular; D018388:Codon, Terminator; D018076:DNA, Complementary / Q000235:genetics; D006801:Humans; D008297:Male; D008854:Microscopy, Electron; D010750:Phosphoproteins / Q000737:chemistry / Q000235:genetics* / Q000276:immunology / Q000378:metabolism*; D011110:Polymorphism, Genetic; D014176:Protein Biosynthesis; D020033:Protein Isoforms / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D051381:Rats; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D013082:Sperm Tail / Q000378:metabolism* / Q000648:ultrastructure; D013091:Spermatogenesis
D000906:Antibodies; C448922:CABYR protein, human; D002135:Calcium-Binding Proteins; D018388:Codon, Terminator; D018076:DNA, Complementary; D010750:Phosphoproteins; D020033:Protein Isoforms; D011994:Recombinant Proteins; D002118:Calcium