Absolute mRNA levels and transcriptional regulation of the mouse testis-specific thioredoxins
Section snippets
Materials and methods
Luciferase reporter constructs. BAC clones RP23-291E22, RP23-332H6, and RP23-332H16 containing mouse Sptrx-1, Sptrx-2, and Sptrx-3 genomic regions, respectively, were obtained from BACPAC Resources Center (http://bacpac.chori.org/). All the Sptrx 5′-flanking regions were amplified by PCR using combinations of the mutagenic primers listed in Supplemental data Table 1. All the PCR products were first cloned into pGEM-Teasy (Promega), verified by sequencing, and inserted into the promoterless
Sptrx mRNA and protein expression levels in GC-spd(ts) cell line
The identification of the cis-acting and trans-acting elements directing the spermatogenic-specific transcription of Sptrx genes is essential to shed more light on their role in sperm physiology. However, a major limitation to this approach is the availability of adequate cell lines that can recapitulate different aspects of spermatogenesis. To our knowledge, GC-spd(ts) is the only spermatogenic mouse cell line available, which is able to undergo meiosis in vitro and develop a primordial tail
Discussion
Regulation of gene expression during spermatogenesis is exerted according to a complex and coordinated scheme where different cis-acting and trans-acting elements converge to either activate, modify or silence the expression of a particular group of specific genes responsible for the formation of a specialized cell as the spermatozoon. Among those gene products participating in the spermatogenic process, we have recently identified three sperm-specific thioredoxins [5]. Since control of redox
Acknowledgments
This work was supported by the Swedish Medical Research Council (Projects 03P-14096, 03X-14041, and 13X-10370), the Åke Wibergs Stiftelse, the Karolinska Institutet, and the Spanish Ministerio de Ciencia y Tecnología (Grant BMC2002-00179). A. Jiménez was supported by a postdoctoral fellowship (EX2003-0390) from the Spanish Ministerio de Educación, Cultura y Deporte. M.-J. Prieto-Álamo and J. Jurado were recipients of postdoctoral contracts (Programa Ramón y Cajal) from the Spanish Ministerio de
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These authors contributed equally to this work.