Cloning, expression and characterization of mouse spermatid specific thioredoxin-1 gene and protein.
Thioredoxins are proteins that participate in different cellular processes via redox-mediated reactions. For humans, we have recently described two novel members of this family that display a male germ cell specific expression pattern, named spermatid specific thioredoxin (Sptrx-1 and Sptrx-2 respectively). We report here the cloning and characterization of the mouse Sptrx-1 gene and protein, which are similar to those described for the human orthologue. The mouse Sptrx-1 open reading frame encodes for a protein of 462 aa composed of an N-terminal repetitive domain of a 15 residue motif followed by a C-terminal domain typical of thioredoxins. The mouse Sptrx-1 gene sequence is interrupted by only one intron of 525 bp located in the 5'-UTR, and using fluorescence in-situ hybridization we have mapped its chromosomal location to 17E1.2-1.3. Northern blot analysis identified the testis as the only tissue expressing mouse Sptrx-1 mRNA, and by in-situ hybridization we found a strong labelling in the testicular seminiferous tubules, mostly in the round spermatids. Affinity purified antibodies against human Sptrx-1 crossreacted well with the mouse protein confirming its expression in seminiferous tubules at the later stages of spermiogenesis. Recombinant mouse Sptrx-1 displayed protein disulphide reducing activity in an enzymatic assay coupled to NADPH and thioredoxin reductase. The availability of the mouse Sptrx-1 gene sequence is the first step towards the generation of knock-out mice, whose characterization will provide significant information regarding the in-vivo function of Sptrx-1 and its possible implication in several sperm anomalies.
Jiménez
Alberto
A
Center for Biotechnology, Department of Biosciences at NOVUM, Karolinska Institutet, S-14157 Huddinge, Sweden.
Oko
Richard
R
Gustafsson
Jan-Ake
JA
Spyrou
Giannis
G
Pelto-Huikko
Markku
M
Miranda-Vizuete
Antonio
A
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D015152:Blotting, Northern; D002874:Chromosome Mapping; D003001:Cloning, Molecular; D008297:Male; D008565:Membrane Proteins / Q000235:genetics* / Q000378:metabolism; D051379:Mice; D008969:Molecular Sequence Data; D009928:Organ Specificity; D012333:RNA, Messenger / Q000378:metabolism; D016415:Sequence Alignment; D013087:Spermatids / Q000378:metabolism*; D013737:Testis / Q000378:metabolism; D013879:Thioredoxins / Q000235:genetics* / Q000378:metabolism
D008565:Membrane Proteins; D012333:RNA, Messenger; C491200:Txndc2 protein, mouse; D013879:Thioredoxins