Dataset Information

Proteomic dissection of dome formation in a mammary cell line: role of tropomyosin-5b and maspin.

ABSTRACT: In this work we extended the study of genes controlling the formation of specific differentiation structures called "domes" formed by the rat mammary adenocarcinoma cell line LA7 under the influence of DMSO. We have reported previously that an interferon-inducible gene, rat-8, and the beta-subunit of the epithelial sodium channel (ENaC) play a fundamental role in this process. Now, we used a proteomic approach to identify proteins differentially expressed either in DMSO-induced LA7 or in 106A10 cells. Two differentially expressed proteins were investigated. The first, tropomyosin-5b, strongly expressed in DMSO-induced LA7 cells, is needed for dome formation because its synthesis inhibition by the antisense RNA technology abolished domes. The second protein, maspin, strongly expressed in the uninduced 106A10 cell line, inhibits dome formation because 106A10 cells, transfected with rat8 cDNA (the function of which is required for the organization of these structures), acquired the ability to develop domes when cultured in presence of an antimaspin antibody. Dome formation in these cultures are accompanied by ENaC beta-subunit expression in the absence of DMSO. Therefore, dome formation requires the expression of tropomyosin-5b, in addition to the ENaC beta-subunit and the rat8 proteins, and is under the negative control of maspin.

SUBMITTER: Zucchi I

PROVIDER: S-EPMC33260 | biostudies-literature | 2001 May

REPOSITORIES: biostudies-literature

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Proteomic dissection of dome formation in a mammary cell line: role of tropomyosin-5b and maspin.

Zucchi I I Bini L L Valaperta R R Ginestra A A Albani D D Susani L L Sanchez J C JC Liberatori S S Magi B B Raggiaschi R R Hochstrasser D F DF Pallini V V Vezzoni P P Dulbecco R R

Proceedings of the National Academy of Sciences of the United States of America 20010501 10

In this work we extended the study of genes controlling the formation of specific differentiation structures called "domes" formed by the rat mammary adenocarcinoma cell line LA7 under the influence of DMSO. We have reported previously that an interferon-inducible gene, rat-8, and the beta-subunit of the epithelial sodium channel (ENaC) play a fundamental role in this process. Now, we used a proteomic approach to identify proteins differentially expressed either in DMSO-induced LA7 or in 106A10 ...[more]

PMID: 11331746

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Project description:Maspin (SerpinB5) is a non-inhibitory serpin (serine protease inhibitor) with very diverse biological activities including regulation of cell adhesion, migration, death, control of gene expression and oxidative stress response. Initially described as a tumor and metastasis suppressor, clinical data brought controversies to the field, as some studies reported no correlation between SerpinB5 expression and prognosis value. These data underscore the importance of understanding SerpinB5 function in a normal physiological context and the molecular mechanism involved. Several SerpinB5 phosphoforms have been detected in different cell lines, but the signaling pathways involved and the biological significance of this post-translational modification in vivo remains to be explored. In this study we investigated SerpinB5 expression, subcellular localization and phosphorylation in different stages of the mouse mammary gland development and the signaling pathway involved. Here we show that SerpinB5 is first detected in late pregnancy, reaches its highest levels in lactation and remains at constant levels during post-lactational regression (involution). Using high resolution isoelectric focusing followed but immunoblot, we found at least 8 different phosphoforms of SerpinB5 during lactation, which decreases steadily at the onset of involution. In order to investigate the signaling pathway involved in SerpinB5 phosphorylation, we took advantage of the non-transformed MCF-10A model system, as we have previously observed SerpinB5 phosphorylation in these cells. We detected basal levels of SerpinB5 phosphorylation in serum- and growth factor-starved cells, which is due to amphiregulin autocrine activity on MCF-10A cells. EGF and TGF alpha, two other EGFR ligands, promote important SerpinB5 phosphorylation. Interestingly, EGF treatment is followed by SerpinB5 nuclear accumulation. Altogether, these data indicate that SerpinB5 expression and phosphorylation are developmentally regulated. In vitro analyses indicate that SerpinB5 phosphorylation is regulated by EGFR ligands, but EGF appears to be the only able to induce SerpinB5 nuclear localization.

Project description:Chromosomal rearrangements that lead to recombination suppression can have a significant impact on speciation, and they are also important for breeding. The regions of recombination suppression in wheat chromosome 5B were identified based on comparisons of the 5B map of a cross between the Chinese Spring (CS) variety of hexaploid wheat and CS-5Bdic (genotype CS with 5B substituted with its homologue from tetraploid Triticum dicoccoides) with several 5B maps of tetraploid and hexaploid wheat. In total, two regions were selected in which recombination suppression occurred in cross CS × CS-5Bdic when compared with other maps: one on the short arm, 5BS_RS, limited by markers BS00009810/BS00022336, and the second on the long arm, 5BL_RS, between markers Ra_c10633_2155 and BS00087043. The regions marked as 5BS_RS and 5BL_RS, with lengths of 5 Mb and 3.6 Mb, respectively, were mined from the 5B pseudomolecule of CS and compared to the homoeologous regions (7.6 and 3.8 Mb, respectively) of the 5B pseudomolecule of Zavitan (T. dicoccoides). It was shown that, in the case of 5BS_RS, the local heterochromatin islands determined by the satellite DNA (119.2) and transposable element arrays, as well as the dissimilarity caused by large insertions/deletions (chromosome rearrangements) between 5BSs aestivum/dicoccoides, are likely the key determinants of recombination suppression in the region. Two major and two minor segments with significant loss of similarity were recognized within the 5BL_RS region. It was shown that the loss of similarity, which can lead to suppression of recombination in the 5BL_RS region, is caused by chromosomal rearrangements, driven by the activity of mobile genetic elements (both DNA transposons and long terminal repeat retrotransposons) and their divergence during evolution. It was noted that the regions marked as 5BS_RS and 5BL_RS are associated with chromosomal rearrangements identified earlier by С-banding analysis of intraspecific polymorphism of tetraploid emmer wheat. The revealed divergence in 5BS_RS and 5BL_RS may be a consequence of interspecific hybridization, plant genetic adaptation, or both.

Dataset Information

Proteomic dissection of dome formation in a mammary cell line: role of tropomyosin-5b and maspin.

Publications

Proteomic dissection of dome formation in a mammary cell line: role of tropomyosin-5b and maspin.

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