scholarly journals Molecular cloning and heterologous expression of an alternatively spliced human Mu class glutathione S-transferase transcript

1993 ◽  
Vol 294 (2) ◽  
pp. 373-380 ◽  
Author(s):  
V L Ross ◽  
P G Board
Keyword(s):  
Alternative Splicing ◽  
Evolutionary Process ◽  
Complete Sequence ◽  
Human Testis ◽  
Cdna Clones ◽  
Glutathione S Transferase ◽  
Testis Cdna Library ◽  
Alternatively Spliced ◽  
Testis Cdna ◽  
Illegitimate Transcription

Two cDNA clones encoding a new Mu class glutathione S-transferase (GST) have been isolated from a human testis cDNA library. Both clones are incomplete and appear to result from alternative splicing. One clone is missing the sequence encoding exon 4 and the other is missing exon 8. The complete sequence of the previously undescribed isoenzyme can be deduced from the two cDNA clones. This is the first report of alternative splicing in a GST transcript and may represent either a novel form of regulation in this multigene family or illegitimate transcription and experimental alternative splicing as part of the evolutionary process. By combining components from each clone a complete cDNA has been constructed and the encoded protein expressed in Escherichia coli. In general, the recombinant enzyme has relatively low activity when compared with all the previously described human Mu class GST isoenzymes.

Identification and characterization of a gene coding a novel isoform of DEAD-box protein

2002 ◽  
Vol 14 (3) ◽  
pp. 185 ◽  
Author(s):  
Lanlan Yin ◽  
JianMin Li ◽  
Hu Zhu ◽  
Min Lin ◽  
Lijun Cheng ◽  
...  
Keyword(s):  
Human Testis ◽  
Microarray Hybridization ◽  
Chain Reaction ◽  
Dead Box ◽  
Gene Coding ◽  
Testis Cdna Library ◽  
Testis Cdna ◽  
Polymerase Chain ◽  
Identification And Characterization

A gene coding a novel isoform of DEAD-box protein named testicular DEAD-box protein (tDbp), presumably involved in testicular function, was identified and characterized. Testicular DEAD-box protein was cloned from a human testis cDNA library. The cDNA microarray hybridization showed that it was expressed at a higher level in adult testis than in embryo testis. Reverse transcription-polymerase chain reaction indicated that tDbp was specifically expressed in testis, but not in some other tissues.

scholarly journals The Chlamydomonas PF6 Locus Encodes a Large Alanine/Proline-Rich Polypeptide That Is Required for Assembly of a Central Pair Projection and Regulates Flagellar Motility

2001 ◽  
Vol 12 (3) ◽  
pp. 739-751 ◽  
Author(s):  
Gerald Rupp ◽  
Eileen O'Toole ◽  
Mary E. Porter
Keyword(s):  
Insertional Mutagenesis ◽  
Expressed Sequence Tag ◽  
Human Testis ◽  
Flagellar Motility ◽  
Central Pair ◽  
Testis Cdna Library ◽  
Motility Mutant ◽  
Testis Cdna ◽  
Temporal And Spatial ◽  
Expressed Sequence

Efficient motility of the eukaryotic flagellum requires precise temporal and spatial control of its constituent dynein motors. The central pair and its associated structures have been implicated as important members of a signal transduction cascade that ultimately regulates dynein arm activity. To identify central pair components involved in this process, we characterized aChlamydomonas motility mutant (pf6-2) obtained by insertional mutagenesis. pf6-2 flagella twitch ineffectively and lack the 1a projection on the C1 microtubule of the central pair. Transformation with constructs containing a full-length, wild-type copy of the PF6 gene rescues the functional, structural, and biochemical defects associated with the pf6 mutation. Sequence analysis indicates that the PF6 gene encodes a large polypeptide that contains numerous alanine-rich, proline-rich, and basic domains and has limited homology to an expressed sequence tag derived from a human testis cDNA library. Biochemical analysis of an epitope-tagged PF6 construct demonstrates that the PF6 polypeptide is an axonemal component that cosediments at 12.6S with several other polypeptides. The PF6 protein appears to be an essential component required for assembly of some of these polypeptides into the C1-1a projection.

scholarly journals Alternative splicing of chicken fibronectin in embryos and in normal and transformed cells.

1987 ◽  
Vol 7 (12) ◽  
pp. 4297-4307 ◽  
Author(s):  
P A Norton ◽  
R O Hynes
Keyword(s):  
Alternative Splicing ◽  
Mrna Level ◽  
Receptor Expression ◽  
Transcriptional Level ◽  
Cdna Clones ◽  
Mrna Levels ◽  
Oncogenic Transformation ◽  
Rous Sarcoma ◽  
Alternatively Spliced ◽  
V Region

To study the alternative splicing of fibronectin during embryogenesis and oncogenic transformation, we isolated cDNA clones of chicken fibronectin. The partial amino acid sequence deduced from sequencing of these clones showed that, overall, chicken fibronectin is approximately 80% identical with mammalian fibronectins. However, two of the three known regions of alternative splicing differed from this average. The V region was significantly more divergent, and RNA from embryonic chicken liver showed a pattern of V exon splicing which was distinct from that seen in human or rat fibronectins. In contrast, the EIIIB segment was very highly conserved (96%). As in mammals, this segment and another (EIIIA) were alternatively spliced in a cell-type-specific fashion. EIIIA+ and EIIIB+ species were almost absent in liver but predominated in total embryo RNA at all times from 2.5 to 11 days postfertilization. We also examined the possible contributions of fibronectin splicing and integrin receptor expression to the loss of fibronectin on oncogenic transformation. We detected little change in fibronectin splicing, other than a slight increase in representation of EIIIB+ species in fibroblasts after transformation by Rous sarcoma virus. It was also established that the overall reduction in fibronectin mRNA level observed after transformation was not accompanied by a decrease in integrin mRNA levels, indicating that fibronectin and integrin receptors are not coordinately regulated at the transcriptional level.

A novel testis protein, RSB-66, interacting with INCA1 (inhibitor of Cdk interacting with cyclin A1)

2008 ◽  
Vol 86 (4) ◽  
pp. 345-351 ◽  
Author(s):  
Xu Chen ◽  
Tinghui Hu ◽  
Gang Liang ◽  
Maojun Yang ◽  
Shudong Zong ◽  
...  
Keyword(s):  
Round Spermatid ◽  
Human Testis ◽  
Amino Acid Residues ◽  
Yeast Two Hybrid ◽  
Cyclin A1 ◽  
Yeast Two Hybrid System ◽  
Testis Cdna Library ◽  
Testis Cdna ◽  
Α Helix ◽  
Two Hybrid

rsb-66 is a novel gene from a suppression subtracted hybridization (SSH) library of round spermatid-specific cDNAs against those of primary spermatocytes. It was found to be specifically expressed in round spermatids. To explore the function of RSB-66, a yeast two-hybrid system was used to screen for potential interacting partners in a human testis cDNA library. HSD45, also known as INCA1 (inhibitor of Cdk interacting with cyclin A1), was identified as one of the positive clones. The interaction between RSB-66 and INCA1 was demonstrated to occur by GST pull down and coimmunoprecipitation. Using immunofluorescence, RSB-66 was found to be specifically expressed in round spermatids, mainly in the cytoplasm. When being transfected into HeLa cells, RSB-66 and INCA1 were found to be co-localized principally in the cytoplasm. The α helix in the RSB-66 C terminal and two amino acid residues (tyr117 and his119) appear to be crucial for its function.

scholarly journals The Drosophila melanogaster tropomyosin II gene produces multiple proteins by use of alternative tissue-specific promoters and alternative splicing.

1988 ◽  
Vol 8 (9) ◽  
pp. 3591-3602 ◽  
Author(s):  
P D Hanke ◽  
R V Storti
Keyword(s):  
Drosophila Melanogaster ◽  
Alternative Splicing ◽  
Gene Evolution ◽  
Cdna Clones ◽  
Developmentally Regulated ◽  
Dna Coding ◽  
Tropomyosin Isoforms ◽  
Alternatively Spliced ◽  
Transcriptional Units ◽  
Regulated Promoters

The structure of the Drosophila melanogaster tropomyosin II (TmII) gene has been determined by DNA sequencing of cDNA clones and the genomic DNA coding for the gene. Two overlapping transcriptional units produce at least four different tropomyosin isoforms. A combination of developmentally regulated promoters and alternative splicing produces both muscle and cytoskeletal tropomyosin isoforms. One promoter is a muscle-specific promoter and produces three different tropomyosin isoforms by alternative splicing of the last three 3' exons. The second promoter has the characteristics of a housekeeping promoter and produces a cytoskeletal tropomyosin isoform. Several internal exons along with a final 3' exon are alternatively spliced in the cytoskeletal transcript. The intron-exon boundaries of the TmII gene are identical to the intron-exon boundaries of all vertebrate tropomyosin genes reported, but are very different from the intron-exon boundaries of the D. melanogaster tropomyosin I gene. The TmII gene is the only reported tropomyosin gene that has two promoters and a quadruple alternative splice choice for the final exon. Models for the mechanism of D. melanogaster tropomyosin gene evolution are discussed.

The Drosophila melanogaster tropomyosin II gene produces multiple proteins by use of alternative tissue-specific promoters and alternative splicing

1988 ◽  
Vol 8 (9) ◽  
pp. 3591-3602
Author(s):  
P D Hanke ◽  
R V Storti
Keyword(s):  
Drosophila Melanogaster ◽  
Alternative Splicing ◽  
Gene Evolution ◽  
Cdna Clones ◽  
Developmentally Regulated ◽  
Dna Coding ◽  
Tropomyosin Isoforms ◽  
Alternatively Spliced ◽  
Transcriptional Units ◽  
Regulated Promoters

The structure of the Drosophila melanogaster tropomyosin II (TmII) gene has been determined by DNA sequencing of cDNA clones and the genomic DNA coding for the gene. Two overlapping transcriptional units produce at least four different tropomyosin isoforms. A combination of developmentally regulated promoters and alternative splicing produces both muscle and cytoskeletal tropomyosin isoforms. One promoter is a muscle-specific promoter and produces three different tropomyosin isoforms by alternative splicing of the last three 3' exons. The second promoter has the characteristics of a housekeeping promoter and produces a cytoskeletal tropomyosin isoform. Several internal exons along with a final 3' exon are alternatively spliced in the cytoskeletal transcript. The intron-exon boundaries of the TmII gene are identical to the intron-exon boundaries of all vertebrate tropomyosin genes reported, but are very different from the intron-exon boundaries of the D. melanogaster tropomyosin I gene. The TmII gene is the only reported tropomyosin gene that has two promoters and a quadruple alternative splice choice for the final exon. Models for the mechanism of D. melanogaster tropomyosin gene evolution are discussed.

Alternative splicing of chicken fibronectin in embryos and in normal and transformed cells

1987 ◽  
Vol 7 (12) ◽  
pp. 4297-4307
Author(s):  
P A Norton ◽  
R O Hynes
Keyword(s):  
Alternative Splicing ◽  
Mrna Level ◽  
Receptor Expression ◽  
Transcriptional Level ◽  
Cdna Clones ◽  
Mrna Levels ◽  
Oncogenic Transformation ◽  
Rous Sarcoma ◽  
Alternatively Spliced ◽  
V Region

To study the alternative splicing of fibronectin during embryogenesis and oncogenic transformation, we isolated cDNA clones of chicken fibronectin. The partial amino acid sequence deduced from sequencing of these clones showed that, overall, chicken fibronectin is approximately 80% identical with mammalian fibronectins. However, two of the three known regions of alternative splicing differed from this average. The V region was significantly more divergent, and RNA from embryonic chicken liver showed a pattern of V exon splicing which was distinct from that seen in human or rat fibronectins. In contrast, the EIIIB segment was very highly conserved (96%). As in mammals, this segment and another (EIIIA) were alternatively spliced in a cell-type-specific fashion. EIIIA+ and EIIIB+ species were almost absent in liver but predominated in total embryo RNA at all times from 2.5 to 11 days postfertilization. We also examined the possible contributions of fibronectin splicing and integrin receptor expression to the loss of fibronectin on oncogenic transformation. We detected little change in fibronectin splicing, other than a slight increase in representation of EIIIB+ species in fibroblasts after transformation by Rous sarcoma virus. It was also established that the overall reduction in fibronectin mRNA level observed after transformation was not accompanied by a decrease in integrin mRNA levels, indicating that fibronectin and integrin receptors are not coordinately regulated at the transcriptional level.

Human testis cDNAs identified by sera from infertile patients: a molecular biological approach to immunocontraceptive development

1994 ◽  
Vol 6 (3) ◽  
pp. 297 ◽  
Author(s):  
ZG Liang ◽  
PA O'Hern ◽  
B Yavetz ◽  
H Yavetz ◽  
E Goldberg
Keyword(s):  
Human Testis ◽  
Vaccine Candidates ◽  
Contraceptive Vaccine ◽  
Immunological Infertility ◽  
Testis Cdna Library ◽  
Human Sera ◽  
Testis Cdna ◽  
Immunologic Infertility ◽  
Infertile Patients ◽  
Molecular Biological Approach

Sera from patients with known or suspected immunological infertility were used to screen a human testis cDNA library. A total of 59 sera detected 38 unique cDNA inserts of which four were testis specific by Northern blot analyses. One of these is a testis-specific isoform of calpastatin. Five additional clones, although not testis specific, were found to be testis abundant. The number and type of clones identified by these human sera suggests a possible aetiology for immunologic infertility. The testis-specific clones will be further characterized to establish their usefulness as contraceptive vaccine candidates.

Su(Ste) Diverged Tandem Repeats in a Y Chromosome of Drosophila melanogaster Are Transcribed and Variously Processed

Genetics ◽  
1998 ◽  
Vol 148 (1) ◽  
pp. 243-249
Author(s):  
Alla I Kalmykova ◽  
Anna A Dobritsa ◽  
Vladimir A Gvozdev
Keyword(s):  
Drosophila Melanogaster ◽  
Rna Processing ◽  
Tandem Repeats ◽  
Antisense Transcription ◽  
Cdna Clones ◽  
Β Subunit ◽  
Splice Sites ◽  
Testis Cdna Library ◽  
Testis Cdna ◽  
Polyadenylation Signals

Abstract We report the organization and transcription of diverged tandemly repeated Y-linked Su(Ste) genes that are considered as suppressors of testis-expressed X-linked-repeated Stellate genes that encode a protein sharing extensive homology with β-subunit of casein kinase 2. Clustering of restriction variants is confirmed. Size variants of Su(Ste) repeats appeared to be nonhomogeneously distributed among the P1 phage clones. Different ways of Su(Ste) RNA processing because of the appearance of new splice sites and polyadenylation signals were detected. The high extent of homology between Stellate and Su(Ste) repeats suggested a possibility of Stellate suppression by antisense transcription of Su(Ste) elements. The detection of only “sense” Su(Ste) cDNAs in testis cDNA library allows us to reject this proposal. The genomic and cDNA clones are shown to be equally diverged. This indicates widespread rather than restricted transcription capacity of these repeats.

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