Genetic modifiers regulating DNA replication and double-strand break repair are associated with differences in mammary tumors in mouse models of Li-Fraumeni syndrome.

Breast cancer is the most common tumor among women with inherited variants in the TP53 tumor suppressor, but onset varies widely suggesting interactions with genetic or environmental factors. Rodent models haploinsufficent for Trp53 also develop a wide variety of malignancies associated with Li-Fraumeni syndrome, but BALB/c mice are uniquely susceptible to mammary tumors and is genetically linked to the Suprmam1 locus on chromosome 7. To define mechanisms that interact with deficiencies in p53 to alter susceptibility to mammary tumors, we fine mapped the Suprmam1 locus in females from an N2 backcross of BALB/cMed and C57BL/6J mice. A major modifier was localized within a 10 cM interval on chromosome 7. The effect of the locus on DNA damage responses was examined in the parental strains and mice that are congenic for C57BL/6J alleles on the BALB/cMed background (SM1-Trp53+/-). The mammary epithelium of C57BL/6J-Trp53+/- females exhibited little radiation-induced apoptosis compared to BALB/cMed-Trp53+/- and SM1-Trp53+/- females indicating that the Suprmam1B6/B6 alleles could not rescue repair of radiation-induced DNA double-strand breaks mostly relying on non-homologous end joining. In contrast, the Suprmam1B6/B6 alleles in SM1-Trp53+/- mice were sufficient to confer the C57BL/6J-Trp53+/- phenotypes in homology-directed repair and replication fork progression. The Suprmam1B6/B6 alleles in SM1-Trp53+/- mice appear to act in trans to regulate a panel of DNA repair and replication genes which lie outside the locus.

Mechano-transduction mediated secretion and uptake of galectin-3 in breast carcinoma cells: implications in the extracellular functions of the lectin.

In the following experiments, we sought to understand the triggering mechanism which propels galectin-3 to be secreted into the extracellular compartment from its intracellular stores in breast carcinoma cells. We also wanted to analyze in greater details the role of galectin-3 in cellular adhesion and spreading. To do this, we made use of two pairs of breast carcinoma cell lines where one of the pair has high expression of galectin-3 and the other low expression of the lectin. We determined that galectin-3 secreted into the conditioned medium of sub-confluent and spread cells in culture was quite low, almost negligible. However, once the cells were detached and rounded up, a mechano-sensing mechanism triggered the rapid secretion of galectin-3 into the conditioned medium. The secretion was constitutive as long as the cells remained detached. Galectin-3 was shown to be actively taken up from the conditioned medium by spreading cells. The cells which express and secrete high levels of galectin-3 adhered and spread much faster on plastic than those with reduced expression. The uptake of galectin-3 according to our data was important in cell spreading because if this process was compromised significantly, cells failed to spread. The data suggested that galectin-3 uptake modulates the adhesion plaques in that cells which express high levels of galectin-3 have thin-dot like plaques that may be suited for rapid adhesion and spreading while cells in which galectin-3 expression is reduced or knocked-down, have thick and elongated plaques which may be suited for a firmer adhesion to the substratum. Recombinant galectin-3 added exogenously reduced the thickness of the adhesion plaques of tumor cells with reduced galectin-3 expression. Taken together, the present data suggest that galectin-3 once externalized, is a powerful modulator of cellular adhesion and spreading in breast carcinoma cells.

The serum glycoprotein fetuin-A promotes Lewis lung carcinoma tumorigenesis via adhesive-dependent and adhesive-independent mechanisms.

Fetuin-A is a serum glycoprotein in the cystatin family associated with the regulation of soft tissue calcification. We tested the role of systemic fetuin in tumor cell growth and metastasis by injecting Lewis lung carcinoma (LLC) cells into fetuin-A null and their wild-type (WT) littermate control C57BL/6 mice via the tail vein, s.c., and intrasplenic routes. In the experimental metastasis assay, the lungs of the WT mice were filled with metastatic nodules, whereas the lungs of the fetuin-A null mutant mice were virtually free of colonies at the end of 2 weeks. Lung colonization responded to the levels of serum fetuin-A in a dose-dependent manner, as observed by the formation of half as many colonies in mice heterozygous for the fetuin-A locus compared with homozygous WT mice and restoration of lung colonization by the administration of purified fetuin-A to fetuin-A-null mice. Serum fetuin-A also influenced the growth of LLC cells injected s.c.: fetuin-A-null mice developed small s.c. tumors only after a substantial delay. Similarly, intrasplenic injection of LLC cells resulted in rapid colonization of the liver with metastasis to the lungs within 2 weeks in the WT but not fetuin-A null mice. To examine the mechanism by which fetuin-A influences LLC colonization and growth, we showed that LLC tumor cells adhere to fetuin-A in a Ca(2+)-dependent fashion, resulting in growth of the tumor cells. These studies support the role of fetuin-A as a major growth promoter in serum that can influence tumor establishment and growth.