Isolation and identification of F-spondin in the boar testis and its production during testis growth.
F-spondin/vascular smooth muscle cell growth-promoting factor (VSGP), purified from the follicular fluid of adult bovine ovaries, has been identified as a promoter of neuronal differentiation and vascular smooth muscle growth. The objectives of the present study were (1) to clarify whether F-spondin is also produced in the testis, which is ontogenically equivalent to the ovary, and (2) to examine whether production of this protein changes with testicular growth. To isolate F-spondin from the testis, testicular homogenates obtained from 8-week-old boars were sequentially subjected to heparin-Sepharose chromatography, diethylaminoethyl (DEAE)-Sepharose chromatography, and reverse-phase high-performance liquid chromatography (RP-HPLC). The isolated protein had a molecular mass of approximately 110 kDa and was cross-reactive with anti-F-spondin antibody by Western blotting. The purified protein was further characterized by amino acid sequence analysis of its internal peptide. The sequence obtained was GEQCNIVPDN VD, and a homology search indicated that the purified protein is a homologue of rat, human, and bovine F-spondin. By fractionation of the same amounts of testis tissue obtained from 1-, 8-, 16-, and 40-week-old boars, we analyzed age-related production of F-spondin in the testis. Western blotting of the fractions obtained from RP-HPLC revealed the presence of a band at approximately 110 kDa, corresponding to F-spondin, in the testes obtained from boars between 1 and 16 weeks old, but this band was not detected at 40 weeks. These results clearly indicate that (1) the porcine testis produces F-spondin and that (2) production of this protein is evident in the immature porcine testis, but not the adult testis.
Ohnuma
Katsuhiko
K
Reproductive Biology Research Unit (Kannondai), National Institute of Agrobiological Science, Ibaraki, Japan.
Kaneko
Hiroyuki
H
Noguchi
Junko
J
Kikuchi
Kazuhiro
K
Ozawa
Manabu
M
Hasegawa
Yoshihisa
Y
D000595:Amino Acid Sequence; D000818:Animals; D002851:Chromatography, High Pressure Liquid; D005260:Female; D015571:Follicular Fluid / Q000502:physiology; D006133:Growth Substances / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D008297:Male; D009419:Nerve Tissue Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D010455:Peptides / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D020539:Sequence Analysis, Protein; D012741:Sexual Maturation; D034421:Sus scrofa; D013737:Testis / Q000254:growth & development* / Q000502:physiology*
D006133:Growth Substances; D009419:Nerve Tissue Proteins; D010455:Peptides