Ere treated with lidocaine and/or bupivacaine. Cell viability, DNA fragmentation, and annexin V immunofluorescence staining were assessed. The effects on apoptosis-related protein expression were investigated by Western blot analysis. The findings were extended to studies in an in vivo xenograft model. RESULTS: Treatment of breast tumor cells with lidocaine and bupivacaine resulted in inhibition of cell viability via induction of apoptosis. The effects were more prominent in MCF-7 cells than in MCF-10A cells. Treatment with local anesthetics induced caspase 7, 8, 9, and poly ADP-ribose polymerase cleavage. The cleavage of caspase 7 and poly ADP-ribose polymerase induced by local anesthetics were effectively blocked by caspase inhibitors. Furthermore, treatment of MCF-7 xenografts with local anesthetics resulted in higher expression of cleaved caspase 7 and an increase in terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL) staining. CONCLUSION: Lidocaine and bupivacaine induce apoptosis of breast tumor cells at clinically relevant concentrations. Our results reveal previously unrecognized beneficial actions of local anesthetics and call for further studies to assess the oncologic advantages of their use during breast cancer surgery....

Liver receptor homolog‐1 (LRH‐1; NR5A2) is an orphan member of the nuclear receptor superfamily, mainly expressed in endoderm‐derived tissues and in the ovary. In ovarian granulosa and luteal cells, LRH‐1 regulates the expression of genes associated with ovarian steroidogenesis. LRH‐1 can be transported to transcriptionally inactive nuclear bodies after conjugation with small ubiquitin‐related modifier (SUMO). In the present study, we investigated the effects of SUMO modification at five lysine residues of LRH‐1 in rat granulosa cells. Lysine 289 could be conjugated with SUMO‐1 in vitro, and the mutation K289R increased transcriptional activity of LRH‐1, suggesting that SUMO conjugation is associated with transcription repression. Coexpression of SUMO‐1 targets LRH‐1 to the dot‐like nuclear bodies, but the effect of lysine mutations on blocking subnuclear localization depended on the cell type. In COS‐7 cells, mutation of either K173 or K289 prevented SUMO‐1‐mediated translocation of LRH‐1 into nuclear bodies and also reduced the conjugation by SUMO‐1, suggesting that K289 and K173 are two important sites involved in SUMO‐1 modification. In granulosa cells, three or more altered lysine residues were required for nucleoplasm retention. This result suggests that multiple lysine residues are targets for SUMO conjugation in vivo and granulosa cells are more sensitive to SUMO‐1‐mediated LRH‐1 localization to nuclear bodies. Nuclear body localization of LRH‐1 was suppressed by forskolin and cholera toxin. Forskolin treatment obviously influences the expression of members involved in the SUMO pathway. The results obtained in the present study suggest that cAMP signaling could change the dynamic process of sumoylation and repress LRH‐1 targeting to nuclear speckles in rat granulosa cells.

SUMO protease SENP1 is elevated in multiple carcinomas including prostate cancer (PCa). SENP1 exhibits carcinogenic properties; it promotes androgen receptor-dependent and -independent cell proliferation, stabilizes HIF1α, increases VEGF, and supports angiogenesis. However, mice expressing an androgen-responsive promoter driven SENP1-transgene (SENP1-Tg) develop high-grade prostatic intraepithelial neoplasia, but not carcinoma. We now show that tumor suppressive PTEN signaling is induced in SENP1-Tg to enhance prostate epithelial cell apoptosis. SENP1 blocks SUMO1-dependent ubiquitylation and degradation of PTEN. In the absence of SENP1, SUMO1-modified PTEN is sequestered in the cytosol, where binding to ubiquitin-E3 ligase WWP2 occurs. Concurrently, WWP2 is also SUMOylated, which potentiates its interaction with PTEN. Thus, SENP1 directs ubiquitin-E3-substrate association to control PTEN stability. PTEN serves as a barrier for SENP1-mediated prostate carcinogenesis as SENP1-Tg mice develop invasive carcinomas only after PTEN reduction. Hence, SENP1 modulates multiple facets of carcinogenesis and may serve as a target specifically for aggressive PTEN-deficient PCa.

Many Toll-like receptors (TLRs) signal through TNF receptor–associated factor 6 (TRAF6) to activate innate immune responses. Here, we show that somatic nuclear autoantigenic sperm protein (sNASP) binds to TRAF6 to prevent TRAF6 autoubiquitination in unstimulated macrophages. Following LPS stimulation, a complex consisting of sNASP, TRAF6, IRAK4, and casein kinase 2 (CK2) is formed. CK2 phosphorylates sNASP at serine 158, allowing sNASP to dissociate from TRAF6. Free TRAF6 is then autoubiquitinated, followed by activation of downstream signaling pathways. In sNasp S158A knockin (S158A-KI) mice, LPS-treated macrophages could not phosphorylate sNASP, which remained bound to TRAF6. S158A-KI mice were more susceptible to sepsis due to a marked reduction in IL-1β, TNF-α, and IFN-γ production accompanied by an inability to clear bacteria and recruit leukocytes. Furthermore, phosphorylation-regulated release of sNASP from TRAF6 is observed following activation of TLR-1, -2, -4, -5, and -6. Thus, sNASP is a negative regulator of TLR signaling to modulate the innate immune response.