The gene for a T lymphocyte triggering factor from African trypanosomes.

An early and essential event in the protective immune response against most viruses and protozoa is the production of interferon-gamma (IFN-gamma). In contrast, during infection with African trypanosomes, protozoan parasites that cause human sleeping sickness, the increased levels of IFN-gamma do not correlate with a protective response. We showed previously that African trypanosomes express a protein called T lymphocyte triggering factor (TLTF), which triggers CD8(+) T lymphocytes to proliferate and to secrete IFN-gamma. Here, we isolate the gene for TLTF and demonstrate that the recombinant version of TLTF specifically induces CD8(+), but not CD4(+), T cells to secrete IFN-gamma. Studies with TLTF fused to the green fluorescent protein show that TLTF is localized to small vesicles that are found primarily at or near the flagellar pocket, the site of secretion in trypanosomes. TLTF is likely to be only the first example of a class of proteins that we designate as trypanokines, i.e., factors secreted by trypanosomes that modulate the cytokine network of the host immune system for the benefit of the parasite.

Isolation and structural analysis of the rat MyoD gene.

We have cloned and determined the nucleotide (nt) sequence of a 6.5-kb genomic DNA fragment containing the rat MyoD gene (encoding a muscle regulatory factor, MyoD). Mouse fibroblasts transfected with this DNA display a high degree of conversion to a muscle phenotype, suggesting that this genomic clone contains sufficient sequence information to allow the production of the rat MyoD protein in these cells. The 6.5-kb genomic fragment contains the complete coding region of MyoD, distributed over three exons, plus 2.3 kb of 5'-noncoding sequence and 1.4 kb of 3'-noncoding sequence. Based on RNase protection assays, the major transcription start point of MyoD is located 210 nt 5' to a methionine start codon and 26 nt 3' to a TAAATA motif which bears similarity to a consensus recognition sequence (TATA) utilized by eukaryotic RNA polymerase II transcription complexes. The high degree of identity between the amino acid sequence of rat MyoD and the MyoD proteins isolated from other vertebrates indicates that this muscle regulatory protein has been evolutionarily conserved.

Inhibition of myogenesis by the H-ras oncogene: implication of a role for protein kinase C.

Expression of the oncogenic form of H-ras p21 in the mouse myogenic cell line, 23A2, blocks myogenesis and inhibits expression of the myogenic regulatory factor gene, MyoD1. Previous studies from a number of laboratories have demonstrated that the activation of ras p21 is associated with changes in phospholipid metabolism that directly, or indirectly, lead to elevated levels of intracellular diacylglycerol and the subsequent activation of protein kinase C (PKC). To assess the importance of PKC activity to the ras-induced inhibition of skeletal myogenesis, we examined the levels of PKC activity associated with the terminal differentiation of wild-type myoblasts and with the differentiation-defective phenotype of 23A2 ras cells. We demonstrate that there is a 50% reduction in PKC activity during normal myogenesis and that PKC activity is required for myoblast fusion, but not for the transcriptional activation of muscle-specific genes. In contrast, we found that the differentiation-defective 23A2 ras cells possess two- to threefold more PKC activity than wild-type myofibers and that reducing the PKC activity in these cultures does not reverse their non-myogenic phenotype. On the other hand, if PKC activity is downregulated in 23A2 cells before the expression of activated ras p21, myogenesis is not inhibited. These results suggest that activated ras p21 relies on a PKC-dependent signal transduction pathway to initiate, but not to sustain, its negative effects on 23A2 skeletal myogenesis and underscore the potential importance of PKC activity to the proper control of skeletal muscle differentiation.

Vasectomy reversal in Nepal.

Data from 157 men in Nepal who had vasectomy reversal are analysed. Most sought reversal within 5 years of vasectomy. Half of the men sought reversal because of the death of a male child, and about one-fourth because of the loss of a female child. Re-marriage was the primary reason for only 10% of the men. Those having reversal because of the loss of a male child were generally younger, and for almost half of this group, the age of their last child at the time of their vasectomy was under 2 years. The results suggest that the demand for reversal could be considerably reduced by more careful screening of the potential vasectomy acceptors.

Fibroblast growth factor and transforming growth factor beta repress transcription of the myogenic regulatory gene MyoD1.

In this report, we demonstrate that myogenic cultures inhibited from differentiating by treatment with fibroblast growth factor or transforming growth factor beta show reduced levels of MyoD1 mRNA. Although this repression may contribute to the inhibition of myogenesis by growth factors, additional regulatory pathways must be affected, since inhibition still occurs in cultures engineered to constitutively express MyoD1 mRNA.

Cooperation of oncogenes in the multistep transformation of established fibroblasts in culture.

The multistage nature of carcinogenesis observed in a variety of systems has been linked experimentally to the sequential activation and subsequent cooperation of oncogene proteins. Cellular transformation resulting from cooperative interactions between activated oncogenes has been described previously for cultured primary cells and certain established cell lines. Our laboratory is using the mouse embryonic cell line C3H10T1/2 to investigate cooperative transformation mediated by the activated human H-ras gene and several nuclear oncogenes from both viral and cellular sources. Oncogene cooperation in C3H10T1/2 is marked by an increase in focus number and an alteration in focus morphology. Although ras cooperates with myc and fos oncogenes to transform C3H10T1/2 fibroblasts, cooperative transformation is not observed in cells similarly co-transfected with the H-ras and E1A oncogenes. This result appears to be due to a growth inhibitory effect of the EIA gene product on C3H10T1/2 cells. The observed cooperation between the H-ras and MC29 viral gag-myc oncogenes has been used along with site-directed mutagenesis of the gag-myc gene to identify two structural regions within the gag-myc protein that mediate ras/myc cooperativity. In addition, the gag-myc mutants generated for these cooperation studies have been used to map the nuclear localization signal of the gag-myc protein and relate the proper cellular location of the protein to its activity in several transformation assays.

Recent changes in the sociodemographic profile of sterilization acceptors in Nepal.

The records of 8056 Nepalese males and 9291 females sterilized from 1979-1983 were analyzed. Compared with data from previous studies (1970-1976), significant decreases occurred in the average age of female acceptors (30.6 vs. 33.1, P less than 0.01) and wives of male acceptors (28.4 vs. 31.7, P less than 0.01). Also, the average number of living children per couple decreased by one child (4.9-3.9, P less than 0.01) for male acceptors and 0.6 (4.8-4.2, P less than 0.01) for female acceptors. Finally in 83.2% of the cases, the operation was performed within 3 years of the last delivery.

PIP: The records of 8056 Nepalese males and 9291 females sterilized from 1979-1983 were analyzed. Compared with data from previous studies (1970-1976), significant decreases occurred in the average age of female acceptors (30.6 vs. 31.7, P0.01) and wives of male acceptors (28.4 vs. 31.7, P0.01). Also, the average number of living children per couple decreased by 1 child (4.9-3.9, P0.01) for male acceptors and 0.6 (4.8--4.2, P0.01) for female acceptors. Finally in 83.2% of the cases, the operation was performed within 3 years of the last delivery. Since 1970-1971 there has been a progressive rise in the % of sterilization acceptors referred by the health worker. For male acceptors there has been nearly a 3-fold increase. The vast majority of both male and female acceptors elected to have their operations within 2 years of the last delivery, regardless of the outcome. It appears that most couples time sterilization to coincide with the cessation of breastfeeding. Care must be taken to guard against the health worker being overly zealous in motivating and mobilizing potential voluntary sterilization contraception candidates.