DNA Synthesis, Methylation and Degradation During Conjugation in \u3cem\u3eTetrahymena thermophila\u3c/em\u3e

We have investigated the timing of DNA synthesis, methylation and degradation during macronuclear development in the ciliate, Tetrahymena thermophila. DNA synthesis was first detected in the anlagen early in macronuclear development, but the majority of DNA synthesis occurred later, after pair separation. Anlagen DNA was first detectably methylated at GATC sites 3–5 hours after its synthesis. Once initiated, de novo methylation was rapid and complete, occurring between 13.5 and 15 hours of conjugation. The level of methylation of GATC sites was constant throughout the remainder of conjugation, and was similar to that in mock-conjugated cells. Degradation of DNA in the old macronucleus and DNA synthesis in the anlagen began at about the same time. Upon pair separation, less than 20% of old macronuclear DNA remained. A small percentage of nucleotides prelabeled prior to conjugation were recycled in the developing analgen

Methylation of Replicating and Nonreplicating DNA in the Ciliate \u3cem\u3eTetrahymena thermophila\u3c/em\u3e

Methylation of adenine in replicating and nonreplicating DNA of the ciliate Tetrahymena thermophila was examined. In growing cells, 87% of the methylation occurred on the newly replicated daughter strand, but methylation was also detectable on the parental strand. Methylation of nonreplicating DNA from starved cells was demonstrated

Site-Specific Methylation of Adenine in the Nuclear Genome of a Eucaryote, \u3cem\u3eTetrahymena thermophila\u3c/em\u3e

DNA in the polyploid macronucleus of the ciliated protozoan Tetrahymena thermophila contains the modified base N6-methyladenine. We identified two GATC sites which are methylated in most or all of the 45 copies of the macronuclear genome. One site is 2 kilobases 5\u27 to the histone H4-I gene, and the other is 5 kilobases 3\u27 to the 73-kilodalton heat shock protein gene. These sites are de novo methylated between 10 and 16 h after initiation of conjugation, during macronuclear anlage development. The methylation states of these two GATC sites and four other unmethylated GATC sites do not change in the DNA of cells cultured under conditions which change the activity of the genes, including logarithmic growth, starvation, and heat shock

Molecular Analysis of N6-Methyladenine Patterns in \u3cem\u3eTetrahymena thermophila\u3c/em\u3e Nuclear DNA

We have cloned two DNA fragments containing 5\u27-GATC-3\u27 sites at which the adenine is methylated in the macronucleus of the ciliate Tetrahymena thermophila. Using these cloned fragments as molecular probes, we analyzed the maintenance of methylation patterns at two partially and two uniformly methylated sites. Our results suggest that a semiconservative copying model for maintenance of methylation is not sufficient to account for the methylation patterns we found during somatic growth of Tetrahymena. Although we detected hemimethylated molecules in macronuclear DNA, they were present in both replicating and nonreplicating DNA. In addition, we observed that a complex methylation pattern including partially methylated sites was maintained during vegetative growth. This required the activity of a methylase capable of recognizing and modifying sites specified by something other than hemimethylation. We suggest that a eucaryotic maintenance methylase may be capable of discriminating between potential methylation sites to ensure the inheritance of methylation patterns

Perceptions of Diet and Physical Activity Among California Hmong Adults and Youths

IntroductionWe conducted a qualitative study to inform the design of a proposed community-wide campaign to promote increased physical activity and fruit and vegetable consumption among low-income Hmong families.MethodsWe held eight focus groups with parents of children aged 5 to 14 years and with youths aged 11 to 14 years, interviews with key informants in several Hmong communities, and interviews with professionals who conducted physical activity and nutrition activities in these communities. Sessions were tape-recorded and transcribed. We organized data using ATLAS.ti software and then analyzed the content.ResultsFindings suggest that physically active lifestyles and dietary patterns emphasizing fresh foods including fruits and vegetables are valued in the Hmong culture and perceived as essential to good health. Barriers to a healthy lifestyle include limited access to safe spaces, time for adequate physical activity, access to land to grow fresh produce, and time for home preparation of food. Low incomes and marketing of unhealthy foods, particularly to children, are also problematic. Information on the healthy aspects of both traditional foods and American foods is needed in accessible formats and delivered through media and trusted community sources.ConclusionLike other Asian groups, the majority of Hmong are first-generation immigrants. An increase in nutrition-related chronic diseases can be prevented by encouraging and reinforcing the maintenance of traditional eating patterns and active lifestyles

Perceptions of diet and physical activity among California Hmong adults and youths.

IntroductionWe conducted a qualitative study to inform the design of a proposed community-wide campaign to promote increased physical activity and fruit and vegetable consumption among low-income Hmong families.MethodsWe held eight focus groups with parents of children aged 5 to 14 years and with youths aged 11 to 14 years, interviews with key informants in several Hmong communities, and interviews with professionals who conducted physical activity and nutrition activities in these communities. Sessions were tape-recorded and transcribed. We organized data using ATLAS.ti software and then analyzed the content.ResultsFindings suggest that physically active lifestyles and dietary patterns emphasizing fresh foods including fruits and vegetables are valued in the Hmong culture and perceived as essential to good health. Barriers to a healthy lifestyle include limited access to safe spaces, time for adequate physical activity, access to land to grow fresh produce, and time for home preparation of food. Low incomes and marketing of unhealthy foods, particularly to children, are also problematic. Information on the healthy aspects of both traditional foods and American foods is needed in accessible formats and delivered through media and trusted community sources.ConclusionLike other Asian groups, the majority of Hmong are first-generation immigrants. An increase in nutrition-related chronic diseases can be prevented by encouraging and reinforcing the maintenance of traditional eating patterns and active lifestyles

Depletion of CD34+ CD4+ cells in bone marrow from HIV-1-infected individuals

AbstractPancytopenia as a consequence of bone marrow abnormalities is commonly seen in HIV-infected individuals. To examine the effect that HIV-1 has on hematopoietic cells, we compared hematopoietic properties of bone marrow samples from HTV+ patients at various stages of disease with bone marrow samples from uninfected donors. While the absolute number of recovered CD34+ cells and the cloning efficiency of these cells did not differ significantly in HIV+ donors, the percentage of CD34+ CD4+ cells was significantly depleted in late-stage HIV+ patients. We observed a direct correlation between the numbers of CD34+ CD4+ cells in the bone marrow and the peripheral CD4 count. Further characterization of the CD34+ CD4+ subpopulation demonstrated that these cells expressed lower levels of HLA-DR on their surface compared with CD34+ CD4- cells, suggesting an immature phenotype. We also found evidence for expression of HIV-1 coreceptors CXCR-4 and CKR-5 message and protein in CD34+ bone marrow cells. While this finding suggested that hematopoietic cells might be susceptible to HIV infection at an early stage of maturation, thus affecting different cell lineages as they matured, we did not find any evidence for infection of HIV in these cells. These data suggest that HIV affects early hematopoietic progenitor cells either directly or indirectly, and in particular CD34+ CD4+ cells. This finding has important implications for disease pathogenesis and for application of gene therapy approaches that use CD34+ hematopoietic cells.Biol Blood Marrow Transplant 1999;5(3):162-72