c-myc in whitefish (Coregonus lavaretus): structure, expression, and insights into possible posttranscriptional regulatory mechanism.

c-myc has a crucial function in growth control, differentiation, and apoptosis of vertebrate cells. Despite the important role of c-myc in mediating the biological effects, studies of c-myc gene expression and factors that control it in organisms other than mammals, such as fish, have been rare. In the current study, we asked whether c-myc mRNA of whitefish, a feasible organism for pollution monitoring in aquatic systems and a model in toxicological research, contains activity sites for regulatory motifs in its 5'- and 3'-UTRs, similar to those found in mammals. We were particularly interested in whether miRNA-34, a known negative regulator of c-myc's in mammals, is able to regulate c-myc in fish. To answer these questions, we determined the mRNA sequence of whitefish c-myc and inferred the structure of the protein that it codes for. We found that the active sites of mRNA and structures of the inferred c-myc protein are similar to those found in mammals and other fish. Remarkably, levels of c-myc mRNA expression were very high in ovaries compared to other tissues of whitefish, thus corroborating previous data in fish. Using bioinformatic searches on c-myc 3'-UTR, we confirmed the presence of two miRNA-34a (miR-34a) response elements. Luciferase reporter assay showed that activity of reporters containing either the miR response elements or entire c-myc 3'-UTR was significantly reduced (p < 0.001) by ectopic expression of miR-34a. Therefore, we further investigated possible involvement of miR-34a in c-myc gene silencing by profiling the expression of both genes in livers of whitefish treated for 8, 24, 48 h with MC-LR, a potent c-myc inducer in mammals. Although the difference was only significant at p = 0.08, the expression of c-myc mRNA in challenged whitefish after 24 h of the treatment was notably higher than that in livers of control fish. Concurrently, we noticed slight but significant up-regulation of miR-34a after 24 and 48 h of the challenge (p < 0.05); however, we found no significant correlation of the c-myc mRNA levels and miR-34a expression. Together, these results suggest that miR-34a might regulate c-myc gene expression in whitefish liver; however, their involvement in MC-LR hepatotoxicity should be clarified in future studies.

Intracellular passage within macrophages affects the trafficking of virulent tubercle bacilli upon reinfection of other macrophages in a serum-dependent manner.

SETTING: The interaction of tubercle bacilli with macrophages is central to understanding of tuberculosis disease. OBJECTIVE: The objective was to determine whether prior passage within macrophages affects the behavior of Mycobacterium tuberculosis (Mtb) upon re-entry into other macrophages. DESIGN: Transmission electron microscopy was used to monitor fusion of bacterial phagosomes with late endosomal/lysosomal compartments using thoria as a fluid phase marker. Two-dimensional polyacrylamide gel electrophoresis was used to study bacterial protein expression within macrophages. RESULTS: H37Rv and BCG expressed novel proteins within macrophages. H37Rv also underwent less fusion after intracellular (IC) (24.2+/-7.7%) than extracellular (XC) (67.4+/-5.5%) passage when the bacteria entered new macrophages in small clusters. These effects were inhibited by serum, and were not observed with H37Ra or BCG bacteria (78.9+/-1.6% fused for all conditions). In addition, vacuoles which contained single bacilli were less likely to acquire markers (26.9+/-2.6%) than those that contained multiple bacilli (77.3+/-2.8%). CONCLUSION: These results indicate that phagolysosomal fusion patterns can be modulated by a variety of factors and that virulent Mtb bacteria may express proteins within macrophages that alter their interaction with these host cells.

Chemotherapy for metastatic melanoma during pregnancy.

Melanoma during pregnancy represents a difficult problem. Although surgery is a definitive therapy for early-stage disease, rapidly progressive metastatic disease in pregnancy requires a series of more difficult decisions. We report the use of combination chemotherapy with tamoxifen, carmustine, dacarbazine, and cisplatin in a patient with metastatic melanoma during the second trimester of pregnancy. The patient received 2 cycles of chemotherapy prior to delivery of a healthy 1520-g baby girl at 30 weeks gestation. Placental pathology, however, revealed malignant melanoma in the intervillous space and melanin pigment granules in villous Hofbauer cells and synctial trophoblasts. This report also reviews the literature, assessing the importance of pregnancy as a prognostic factor, the effects of metastasis on the products of conception, and the use of chemotherapy in pregnancy. We conclude that combination chemotherapy can be administered in the second trimester of pregnancy for the treatment of rapidly progressive melanoma without interfering with the successful maturation and delivery of an infant of 30 weeks gestation.