Abstract
In the postnatal testis, the c-kit transmembrane tyrosine-kinase receptor is expressed in type A spermatogonia, and its transcription ceases at the meiotic phase of spermatogenesis. Alternative, shorter c-kit transcripts are expressed in post-meiotic germ cells. These transcripts should encode a truncated version of the c-kit protein, lacking the extracellular, the transmembrane and part of the intracellular tyrosine-kinase domains. The 5' end of the alternative c-kit transcripts maps within an intron of the mouse c-kit gene. We now show that this intron contains a promoter active in nuclear extracts of round spermatids, and that two discrete sequences upstream of the transcriptional start site bind spermatid-specific nuclear factors. Deletion of both these sequences abolishes activity of the promoter in vitro. We have also established that this promoter is functional in vivo, in a tissue- and cell-specific fashion, since intronic sequences drive the expression of the E, coli lacZ reporter gene in transgenic mice specifically in the testis. Transgene expression is confined to haploid germ cells of seminiferous tubules, starting from spermatids at step 9, and disappearing at step 13, indicating that a cryptic promoter within the 16(th) intron of the mouse c-kit gene is active in a short temporal window at the end of the transcriptional phase of spermiogenesis. In agreement with these data, western blot experiments using an antibody directed against the carboxy-terminal portion of the mouse c-kit protein showed that a polypeptide, of the size predicted by the open reading frame of the spermatid-specific c-kit cDNA, accumulates in the latest stages of spermatogenesis and in epididymal spermatozoa. An immunoreactive protein of the same size can be produced in both eukaryotic and prokaryotic artificial expression systems.
| Original language | English |
|---|---|
| Pages (from-to) | 1291-1302 |
| Number of pages | 12 |
| Journal | Development |
| Volume | 122 |
| Issue number | 4 |
| Publication status | Published - Apr 1996 |
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Keywords
- c-kit
- Cell-specific promoter
- Haploid-specific transcription
- Spermatogenesis
- Transgenic mice
- Tyrosine-kinase-independent function
ASJC Scopus subject areas
- Anatomy
- Cell Biology
Cite this
A cell- and developmental stage-specific promoter drives the expression of a truncated c-kit protein during mouse spermatid elongation. / Albanesi, Cristina; Geremia, Raffaele; Giorgio, Marco; Dolci, Susanna; Sette, Claudio; Rossi, Pellegrino.
In: Development, Vol. 122, No. 4, 04.1996, p. 1291-1302.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - A cell- and developmental stage-specific promoter drives the expression of a truncated c-kit protein during mouse spermatid elongation
AU - Albanesi, Cristina
AU - Geremia, Raffaele
AU - Giorgio, Marco
AU - Dolci, Susanna
AU - Sette, Claudio
AU - Rossi, Pellegrino
PY - 1996/4
Y1 - 1996/4
N2 - In the postnatal testis, the c-kit transmembrane tyrosine-kinase receptor is expressed in type A spermatogonia, and its transcription ceases at the meiotic phase of spermatogenesis. Alternative, shorter c-kit transcripts are expressed in post-meiotic germ cells. These transcripts should encode a truncated version of the c-kit protein, lacking the extracellular, the transmembrane and part of the intracellular tyrosine-kinase domains. The 5' end of the alternative c-kit transcripts maps within an intron of the mouse c-kit gene. We now show that this intron contains a promoter active in nuclear extracts of round spermatids, and that two discrete sequences upstream of the transcriptional start site bind spermatid-specific nuclear factors. Deletion of both these sequences abolishes activity of the promoter in vitro. We have also established that this promoter is functional in vivo, in a tissue- and cell-specific fashion, since intronic sequences drive the expression of the E, coli lacZ reporter gene in transgenic mice specifically in the testis. Transgene expression is confined to haploid germ cells of seminiferous tubules, starting from spermatids at step 9, and disappearing at step 13, indicating that a cryptic promoter within the 16(th) intron of the mouse c-kit gene is active in a short temporal window at the end of the transcriptional phase of spermiogenesis. In agreement with these data, western blot experiments using an antibody directed against the carboxy-terminal portion of the mouse c-kit protein showed that a polypeptide, of the size predicted by the open reading frame of the spermatid-specific c-kit cDNA, accumulates in the latest stages of spermatogenesis and in epididymal spermatozoa. An immunoreactive protein of the same size can be produced in both eukaryotic and prokaryotic artificial expression systems.
AB - In the postnatal testis, the c-kit transmembrane tyrosine-kinase receptor is expressed in type A spermatogonia, and its transcription ceases at the meiotic phase of spermatogenesis. Alternative, shorter c-kit transcripts are expressed in post-meiotic germ cells. These transcripts should encode a truncated version of the c-kit protein, lacking the extracellular, the transmembrane and part of the intracellular tyrosine-kinase domains. The 5' end of the alternative c-kit transcripts maps within an intron of the mouse c-kit gene. We now show that this intron contains a promoter active in nuclear extracts of round spermatids, and that two discrete sequences upstream of the transcriptional start site bind spermatid-specific nuclear factors. Deletion of both these sequences abolishes activity of the promoter in vitro. We have also established that this promoter is functional in vivo, in a tissue- and cell-specific fashion, since intronic sequences drive the expression of the E, coli lacZ reporter gene in transgenic mice specifically in the testis. Transgene expression is confined to haploid germ cells of seminiferous tubules, starting from spermatids at step 9, and disappearing at step 13, indicating that a cryptic promoter within the 16(th) intron of the mouse c-kit gene is active in a short temporal window at the end of the transcriptional phase of spermiogenesis. In agreement with these data, western blot experiments using an antibody directed against the carboxy-terminal portion of the mouse c-kit protein showed that a polypeptide, of the size predicted by the open reading frame of the spermatid-specific c-kit cDNA, accumulates in the latest stages of spermatogenesis and in epididymal spermatozoa. An immunoreactive protein of the same size can be produced in both eukaryotic and prokaryotic artificial expression systems.
KW - c-kit
KW - Cell-specific promoter
KW - Haploid-specific transcription
KW - Spermatogenesis
KW - Transgenic mice
KW - Tyrosine-kinase-independent function
UR - http://www.scopus.com/inward/record.url?scp=0030002951&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=0030002951&partnerID=8YFLogxK
M3 - Article
C2 - 8620856
AN - SCOPUS:0030002951
VL - 122
SP - 1291
EP - 1302
JO - Development (Cambridge)
JF - Development (Cambridge)
SN - 0950-1991
IS - 4
ER -