Capacity for protein synthesis following heat stimulus of Drosophila associates with heat tolerance but does not underlie the latitudinal tolerance cline

Across populations of Drosophila melanogaster along the Australian eastern coastline latitudinal clines occur in both heat-knockdown tolerance and hardened heat-knockdown tolerance – low latitude tropical populations being more tolerant. A latitudinal cline also occurs for rates of total protein synthesis following a mild heat stress, with tropical populations having higher rates. Since the control of protein synthesis following heat stress is an important component of the cellular heat-shock response, we hypothesised that the higher rates of synthesis that follow a heat stimulus lead to higher knockdown tolerance and underpins the cline. However, levels of heat-stimulated total protein synthesis have been negatively related to heat-hardening capacity, a somewhat conflicting result. Here we examine the relationship between these physiological and adaptive traits in a set of 40 family lines derived from a hybrid laboratory population established by crossing populations from either end of the latitudinal transect. Among these lines high levels of heat-stimulated total protein synthesis were associated with both low basal and low heat-hardened adult knockdown time, confirming the importance of a negative relationship between protein synthesis and thermal tolerance. This result, when considered along with the directions of the latitudinal clines in protein synthesis and tolerance, suggests that variation in rates of heat-stimulated total protein synthesis is not a factor contributing to the latitudinal cline in heat tolerance. Given the robustness of this negative relationship we discuss possible explanations and future experiments to elucidate how the cellular heat stress response might facilitate increased knockdown tolerance.     

Identification of an NF-kappa B binding site in the bovine leukemia virus promoter.

Although the mechanism by which bovine leukemia virus (BLV) induces neoplastic transformation of the host B cells is unknown, it is likely that critical interactions between cellular DNA-binding proteins and the virus are involved. We have used DNase I protection (footprinting) assays to construct a map of protein-DNA interactions on the 5' long terminal repeat of BLV. In addition to the three cyclic AMP response elements previously reported, we have also found an NF-kappa B binding site between -118 and -70 nucleotides upstream of the RNA start site. This site binds several members of the kappa B family of proteins, including p49, p50, and p65, in both footprint and electrophoretic mobility shift assays and functions as an enhancer element when inserted upstream of the chloramphenicol acetyltransferase gene. NF-kappa B may be a critical nuclear binding protein that regulates both viral replication and key cellular genes in BLV-infected B cells.     

Macroinvertebrates as bioindicators of water quality in the Mkondoa River,Tanzania, in an agricultural area

The suitability of using macroinvertebrates as bioindicators of stream water quality was tested in the Mkondoa River in an agricultural area at Kilosa, using the rapid bioassessment protocol. The family biotic index (FBI) showed marked variation in water quality along the stream from values ranging from 4.1 to 5.0 in the upstream reaches, indicating good water quality, 5.3 to 5.5 in the mid-reaches and 6.0 to 6.5 in the lower reaches. The water quality index (WQI) indicated that water quality was fair (77 ± 0.98) in the upstream reach of the Mkondoa, marginal (55 ± 0.86) in the midstream reach and poor (33 ± 0.45) in the downstream reach. There were significant relationships between biological oxygen demand and dissolved oxygen and the occurrence of specific taxa, mainly Chironomus and Caenis. Significant changes in macroinvertebrate abundance were mostly related to changes in water quality. As in other parts of the world, macroinvertebrate communities proved to be good biological indicators of water quality and they should be used as bioindicators in long-term monitoring of this river.     

Myosin light-chain expression during avian muscle development

Monoclonal antibodies to adult chicken myosin light chains were generated and used to quantitate the types of myosin light-chain (MLC) isoforms expressed during development of the pectoralis major (PM), anterior latissimus dorsi (ALD), and medial adductor (MA) muscles of the chicken. These are muscles which, in the adult, are composed predominantly of fast, slow, and a mixture of fiber types, respectively. Three distinct phases of MLC expression characterized the development of the PM and MA muscles. The first identifiable pase occurred during the period of 5-7 d of incubation in ovo. Extracts of muscles from the pectoral region (which included the presumptive PM muscle) contained only fast MLC isoforms. This period of exclusive fast light-chain synthesis was followed by a phase (8- 12 d of incubation in ovo) in which coexpression of both fast and slow MLC isoforms was apparent in both PM and MA muscles. During the period, the composition of both fast and slow MLC isoforms in the PM and MA muscles was identical. Beginning at day 12 in ovo, the ALD was also subjected to immunochemical analyses. The proportion of fast and slow MLCs in this muscle at day 12 was similar to that present in the other muscles studied. The third development phase of MLC expression began at approximately 12 d of incubation in ovo and encompassed the transition in MLC composition to the isoform patterns incubation in ovo and encompassed the transition in MLC composition to the isoform patterns typical of adult muscle. During this period, the relative proportion of slow MLC rose in both the MA and ALD and fell in the PM. By day 16, the third fast light chain, LC(3f), was apparent in extracts of both the PM and MA. These results show that there is a developmental progression in the expression of MLC in the two avian muscles studied from day 5 in ovo; first, only fast MLCs are accumulated, then both fast and slow MLC isoforms are expressed. Only during the latter third of development in ovo is the final MLC isoform pattern characteristic of a particular muscle type expressed.     

Pathogenicity of molecularly cloned bovine leukemia virus.

To delineate the mechanisms of bovine leukemia virus (BLV) pathogenesis, four full-length BLV clones, 1, 8, 9, and 13, derived from the transformed cell line FLK-BLV and a clone construct, pBLV913, were introduced into bovine spleen cells by microinjection. Microinjected cells exhibited cytopathic effects and produced BLV p24 and gp51 antigens and infectious virus. The construct, pBLV913, was selected for infection of two sheep by inoculation of microinjected cells. After 15 months, peripheral blood mononuclear cells from these sheep served as inocula for the transfer of infection to four additional sheep. All six infected sheep seroconverted to BLV and had detectable BLV DNA in peripheral blood mononuclear cells after amplification by polymerase chain reaction. Four of the six sheep developed altered B/T-lymphocyte ratios between 33 and 53 months postinfection. One sheep died of unrelated causes, and one remained hematologically normal. Two of the affected sheep developed B lymphocytosis comparable to that observed in animals inoculated with peripheral blood mononuclear cells from BLV-infected cattle. This expanded B-lymphocyte population was characterized by elevated expression of B-cell surface markers, spontaneous blastogenesis, virus expression in vitro, and increased, polyclonally integrated provirus. One of these two sheep developed lymphocytic leukemia-lymphoma at 57 months postinfection. Leukemic cells had the same phenotype and harbored a single, monoclonally integrated provirus but produced no virus after in vitro cultivation. The range in clinical response to in vivo infection with cloned BLV suggests an important role for host immune response in the progression of virus replication and pathogenesis.     

The effect of cell synchronization upon the detection of T and B lymphoid cell receptors on two continuous lymphoid cell lines.

In the present study, the effect of the cell synchronization on the detection of T and B cell surface markers of two continuous lines of lymphoid cells (FL-74 and CT45-S) was examined. Suspension cultures were synchronized by deprivation of isoleucine and surface markers were quantitated by T rosette formation with guinea pig erythrocytes (E) and B rosette formation with an erythrocyte-antibody-complement (EAC) complex. After 24 hr, cells were resuspended in complete culture medium. Virtually 100% of FL-74 cells expressed the T cell marker at time 0, with a progressive decline to 80% at saturation density. A bell-shaped curve for expression of the EAC marker on CT45-S cells was seen with maximum expression in the logarithmic phase of the growth cycle. Spent culture medium was examined for the presence of free soluble receptor. Preincubation of E and EAC in appropriate old medium resulted in 42% inhibition of E rosettes and 42% inhibition of EAC rosettes with FL-74 and CT45-S cells, respectively. Thus quantitation of lymphocyte subpopulations as B, T or null cells with these cellular markers may be influenced by the age of the cell examined, phase of the cell cycle and the amount of free receptor present in the surrounding medium.     

The effect of cell synchronization upon the detection of T and B lymphoid cell receptors on two continuous lymphoid cell lines

Summary In the present study, the effect of the cell synchronization on the detection of T and B cell surface markers of two continuous lines of lymphoid cells (FL-74 and CT45-S) was examined. Suspension cultures were synchronized by deprivation of isoleucine and surface markers were quantitated by T rosette formation with guinea pig erythrocytes (E) and B rosette formation with an erythrocyte-antibody-complement (EAC) complex. After 24 hr, cells were resuspended in complete culture medium. Virtually 100% of FL-74 cells expressed the T cell marker at time 0, with a progressive decline to 80% at saturation density. A bell-shaped curve for expression of the EAC marker on CT45-S cells was seen with maximum expression in the logarithmic phase of the growth cycle. Spent culture medium was examined for the presence of free soluble receptor. Preincubation of E and EAC in appropriate old medium resulted in 42% inhibition of E rosettes and 42% inhibition of EAC rosettes with FL-74 and CT45-S cells, respectively. Thus quantitation of lymphocyte subpopulations as B, T or null cells with these cellular markers may be influenced by the age of the cell examined, phase of the cell cycle and the amount of free receptor present in the surrounding medium. This research was supported in part by contract NO1 CP 5-3571 with the Virus Cancer Program of the NCI, NIH, PHS grant no. 2 RO1 A1-09022-07, Allergy and Infectious Diseases NIH, PHS and The State of Ohio Canine Research Funds.