Dataset Information

Proteome analysis of testis from infertile protein C inhibitor deficient mice reveals predominant changes in serpin processing and prostaglandin metabolism

ABSTRACT: Serine protease inhibitors have therapeutic potential in a variety of pathogenic processes, ranging from thrombosis, altered immune response to liver cirrhosis. To investigate the physiological effects of protein C inhibitor (PCI, serpinA5) this gene was inactivated in a mouse model with the resulting phenotype of male infertility. In the actual report 2D-DIGE analysis was utilized to investigate the molecular mechanisms for PCI in male reproduction. Comparing the testes proteome of three PCI knockout-mice with the three wild-types- demonstrated similar pattern with the exception of a massive upregulation of prostaglandin reductase 1 (10-fold; P < 0.002) and complete shifts in the molecular weights of serpinA1C and serpinA3K. All these PCI-dependent proteome changes were immunologically verified. Unbiased proteome analysis indicated that the inactivation of serpinA5 strongly influenced both the protein species pattern of other A-clade serpins as well as the prostaglandin metabolism in the testes.

INSTRUMENT(S): LTQ Orbitrap Velos, LTQ Velos

ORGANISM(S): Mus Musculus (mouse)

TISSUE(S): Testis

SUBMITTER: Goran Mitulović

LAB HEAD: Maria Zellner

PROVIDER: PXD002944 | Pride | 2015-12-07

REPOSITORIES: Pride

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Publications

Proteome analysis of testis from infertile protein C inhibitor-deficient mice reveals novel changes in serpin processing and prostaglandin metabolism.

Yang Hanjiang H Wahlmüller Felix Christof FC Uhrin Pavel P Baumgartner Roland R Mitulovic Goran G Sarg Bettina B Geiger Margarethe M Zellner Maria M

Electrophoresis 20151020 21-22

Serine protease inhibitors (serpin) have therapeutic potential in a variety of pathogenic processes, ranging from thrombosis and altered immune response to liver cirrhosis. To investigate the physiological effects of protein C inhibitor (PCI, serpinA5), its gene was inactivated in a mouse model, resulting in male infertility. In the present report, 2D differential gel electrophoresis was utilized to investigate the molecular mechanisms for PCI involvement in male reproduction. Comparing the test ...[more]

PMID: 26332751

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Project description:The screening of a cDNA derived expression library of Campylobacter jejuni NCTC 11168 expressed in E. coli using a fusion construct and specific HaloTag interaction to a modified surface is shown. 1536 different clones were screened including positive (hisJ, cjaA, peb1a) and negative (argC, pyrC, gapA) reference proteins. The goal of the screening was to identify potential novel immunogenic proteins from C. jejuni by selecting clones showing a high signal intensity in comparison to the known antigens used as positive markers. Afterwards, the most promising clones were sequenced to identify the gene and corresponding protein, and these proteins were then investigated further. Consequently, 22 novel immunogenic proteins could be identified. In total, 1536 (4 x 384) different lysates were spotted on different microarray slides. Each slide contained 3600 distinct spots, separated into two compartments of 1800 spots each. Each compartment comprised quadruplicate replicates of each sample lysate with controls being analysed with more replicates. As controls we used: hisJ, cjaA and peb1 (3 x 40 replicates) as positive reference proteins, as they have been described as immunogenic before; argC and pyrC (2 x 40 replicates) as negative reference proteins; two sets of E. coli cell lysates without fusion proteins expressed, namely Acella electrocompetent cells (40 replicates) and KRX (32 replicates); and a buffer control (24 replicates). Therefore, each set of replicate slides contained 376 different samples and 8 controls. Each screening was performed with three replicate slides. Consequently, a total number of 12 slides were screened (4 sets of samples x 3 replicates each). For identification, rabbit polyclonal antibody to C. jejuni (Acris AP24002PU-N) as primary and goat polyclonal to rabbit IgG conjugated with ChromeoTM-546 (Abcam ab60317) as secondary antibody were used. The top compartment was incubated first with primary antibody, while the bottom compartment was incubated with PBS at the same time. Afterwards, both compartments were incubated with secondary antibody.

Dataset Information

Proteome analysis of testis from infertile protein C inhibitor deficient mice reveals predominant changes in serpin processing and prostaglandin metabolism

Publications

Proteome analysis of testis from infertile protein C inhibitor-deficient mice reveals novel changes in serpin processing and prostaglandin metabolism.

Similar Datasets

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