Influence of plasma calcium and vitamin D on bone collagen. Effects on lysine hydroxylation and crosslink formation.

In chick bone collagen the degree of lysine hydroxylation and the magnitude of the crosslink ratio dihydroxylysinonorleucine/hydroxylysinonorleucine were both found to be inversely related to the concentration of plasma calcium. Lysine hydroxylation was also affected by a second factor related to vitamin D status.     

Approaches to sequence analysis of 125I-labeled RNA.

A method is described for the initial steps of sequence analysis of RNase T1-and pancreatic RN-ase-resistant oligonucleotides of RNA containing cytidylate residues labeled in vitro with 125I. In many cases an oligonucleotide sequence can be deduced from a consideration of (i) its relative position in the two-dimensional fingerprint (with DEAE thin layer homochromatographic second dimension), (ii) its electrophoretic mobility on DEAE paper at pH 1.9, and (iii) identification of its products of further enzymatic digestion by comparison with a set of marker oligonucleotides. Additional methods including analysis of oligonucleotides following chemical blocking of uridylate residues with CMCT and analysis of products of incomplete enzymatic digestion are also discussed.     

Incorporation of C-photosynthate into major chemical fractions of source and sink leaves of cottonwood

The incorporation and distribution of photosynthetically fixed ¹⁴CO₂ was followed for 48 hours in a recently matured source leaf (LPI 7) and in young expanding source and sink leaves (LPI 4) of cottonwood (Populus deltoides Bartr.). The major chemical constituents of leaf laminae and petioles were separated by sequential solvent extractions and enzyme hydrolyses. Two hours after labeling, about 80% of the ¹⁴C was found in water-alcohol-soluble constituents in the mature source lamina as compared to about 45% in those of the young expanding leaf. In both mature and expanding source leaves the water-alcohol-soluble constituents decreased while the CHCl₃-soluble and -insoluble compounds increased with time. After 48 hours, 7 and 37% of the total ¹⁴C was recovered from structural carbohydrates and from protein + CHCl₃-soluble fractions, respectively, in the mature source leaf; and 4 and 65%, respectively, in the young source leaf. When the distribution of ¹⁴C among major chemical fractions was calculated on per cent dpm/mg basis, the data showed that a young sink leaf incorporated over twice as much ¹⁴C into structural carbohydrates as a young source leaf (11% versus 4%). However, when calculated on an absolute dpm/mg basis, activity in this fraction of the young source leaf exceeded that in the sink leaf by a ratio of about 11:1 (9528 versus 845 dpm/mg). Thus, most of the material for synthesis of structural carbohydrates was derived from in situ photosynthate.     

Temperature Effects on Development of Meloidogyne Enterolobii and M. Floridensis

Meloidogyne enterolobii and M. floridensis are virulent species that can overcome root-knot nematode resistance in economically important crops. Our objectives were to determine the effects of temperature on the infectivity of second-stage juveniles (J2) of these two species and determine differences in duration and thermal-time requirements (degree-days [DD]) to complete their developmental cycle. Florida isolates of M. enterolobii and M. floridensis were compared to M. incognita race 3. Tomato cv. BHN 589 seedlings following inoculation were placed in growth chambers set at constant temperatures of 25°C, and 30°C, and alternating temperatures of 30°C to 25°C (day–night). Root infection by the three nematode species was higher at 30°C than at 25°C, and intermediate at 30°C to 25°C, with 33%, 15%, and 24% infection rates, respectively. There was no difference, however, in the percentages of J2 that infected roots among species at each temperature. Developmental time from infective J2 to reproductive stage for the three species was shorter at 30°C than at 25°C, and 30°C to 25°C. The shortest time and DD to egg production for the three species were 13 days after inoculation (DAI) and 285.7 DD, respectively. During the experimental timeframe of 29 d, a single generation was completed at 30°C for all three species, whereas only M. floridensis completed a generation at 30°C to 25°C. The number of days and accumulated DD for completing the life cycle (from J2 to J2) were 23 d and 506.9 DD for M. enterolobii, and 25 d and 552.3 DD for M. floridensis and M. incognita, respectively. Exposure to lower (25°C) and intermediate temperatures (30°C to 25°C) decreased root penetration and slowed the developmental cycle of M. enterolobii and M. floridensis compared with 30°C.     

Methylated DNA/RNA in Body Fluids as Biomarkers for Lung Cancer

DNA/RNA methylation plays an important role in lung cancer initiation and progression. Liquid biopsy makes use of cells, nucleotides and proteins released from tumor cells into body fluids to help with cancer diagnosis and prognosis. Methylation of circulating tumor DNA (ctDNA) has gained increasing attention as biomarkers for lung cancer. Here we briefly introduce the biological basis and detection method of ctDNA methylation, and review various applications of methylated DNA in body fluids in lung cancer screening, diagnosis, prognosis, monitoring and treatment prediction. We also discuss the emerging role of RNA methylation as biomarkers for cancer.     

Disparities in distribution of COVID-19 vaccines across US counties: A geographic information system–based cross-sectional study

BackgroundThe US Centers for Disease Control and Prevention has repeatedly called for Coronavirus Disease 2019 (COVID-19) vaccine equity. The objective our study was to measure equity in the early distribution of COVID-19 vaccines to healthcare facilities across the US. Specifically, we tested whether the likelihood of a healthcare facility administering COVID-19 vaccines in May 2021 differed by county-level racial composition and degree of urbanicity.Methods and findingsThe outcome was whether an eligible vaccination facility actually administered COVID-19 vaccines as of May 2021, and was defined by spatially matching locations of eligible and actual COVID-19 vaccine administration locations. The outcome was regressed against county-level measures for racial/ethnic composition, urbanicity, income, social vulnerability index, COVID-19 mortality, 2020 election results, and availability of nontraditional vaccination locations using generalized estimating equations.Across the US, 61.4% of eligible healthcare facilities and 76.0% of eligible pharmacies provided COVID-19 vaccinations as of May 2021. Facilities in counties with >42.2% non-Hispanic Black population (i.e., > 95th county percentile of Black race composition) were less likely to serve as COVID-19 vaccine administration locations compared to facilities in counties with <12.5% non-Hispanic Black population (i.e., lower than US average), with OR 0.83; 95% CI, 0.70 to 0.98, p = 0.030. Location of a facility in a rural county (OR 0.82; 95% CI, 0.75 to 0.90, p < 0.001, versus metropolitan county) or in a county in the top quintile of COVID-19 mortality (OR 0.83; 95% CI, 0.75 to 0.93, p = 0.001, versus bottom 4 quintiles) was associated with decreased odds of serving as a COVID-19 vaccine administration location.There was a significant interaction of urbanicity and racial/ethnic composition: In metropolitan counties, facilities in counties with >42.2% non-Hispanic Black population (i.e., >95th county percentile of Black race composition) had 32% (95% CI 14% to 47%, p = 0.001) lower odds of serving as COVID administration facility compared to facilities in counties with below US average Black population. This association between Black composition and odds of a facility serving as vaccine administration facility was not observed in rural or suburban counties. In rural counties, facilities in counties with above US average Hispanic population had 26% (95% CI 11% to 38%, p = 0.002) lower odds of serving as vaccine administration facility compared to facilities in counties with below US average Hispanic population. This association between Hispanic ethnicity and odds of a facility serving as vaccine administration facility was not observed in metropolitan or suburban counties.Our analyses did not include nontraditional vaccination sites and are based on data as of May 2021, thus they represent the early distribution of COVID-19 vaccines. Our results based on this cross-sectional analysis may not be generalizable to later phases of the COVID-19 vaccine distribution process.ConclusionsHealthcare facilities in counties with higher Black composition, in rural areas, and in hardest-hit communities were less likely to serve as COVID-19 vaccine administration locations in May 2021. The lower uptake of COVID-19 vaccinations among minority populations and rural areas has been attributed to vaccine hesitancy; however, decreased access to vaccination sites may be an additional overlooked barrier.

Inmaculada Hernandez and colleagues investigate the disparities in early-phase distribution of COVID-19 Vaccines across U.S. Counties.     

Rat cholesterol side-chain cleavage enzyme (P-450scc): use of a cDNA probe to study the hormonal regulation of P-450scc mRNA levels in ovarian granulosa cells

A rat ovarian cDNA library was constructed and screened by differential colony hybridization to detect cDNA clones specific for mRNA induced by follicle-stimulating hormone (FSH). The cDNA clone which demonstrated the greatest degree of induction contained a 766-bp insert which was characterized and sequenced. We conclude that this cDNA is specific for the rat gene coding for cholesterol side-chain cleavage enzyme (P-450scc) by virtue of nucleotide sequence homology to the bovine and human P-450scc cDNA sequences. Southern blotting of rat genomic DNA suggests the presence of a single P-450scc gene. Northern blot analysis indicates that P-450scc mRNA is present in steroidogenic tissues (ovary, adrenal, testis), but not in brain, kidney, liver, lung, or heart. The rat P-450scc mRNA is induced by FSH or pregnant mare's serum gonadotropin in ovaries of estrogen-treated immature rats in vivo. In cultured granulosa cells, estradiol treatment alone did not increase P-450scc mRNA levels, but in combination with FSH or 8-Br-cAMP resulted in three- to four-fold increase in this mRNA.     

Human ERCC5 cDNA-cosmid complementation for excision repair and bipartite amino acid domains conserved with RAD proteins of Saccharomyces cerevisiae and Schizosaccharomyces pombe.

Several human genes related to DNA excision repair (ER) have been isolated via ER cross-species complementation (ERCC) of UV-sensitive CHO cells. We have now isolated and characterized cDNAs for the human ERCC5 gene that complement CHO UV135 cells. The ERCC5 mRNA size is about 4.6 kb. Our available cDNA clones are partial length, and no single clone was active for UV135 complementation. When cDNAs were mixed pairwise with a cosmid clone containing an overlapping 5'-end segment of the ERCC5 gene, DNA transfer produced UV-resistant colonies with 60 to 95% correction of UV resistance relative to either a genomic ERCC5 DNA transformant or the CHO AA8 progenitor cells. cDNA-cosmid transformants regained intermediate levels (20 to 45%) of ER-dependent reactivation of a UV-damaged pSVCATgpt reporter plasmid. Our evidence strongly implicates an in situ recombination mechanism in cDNA-cosmid complementation for ER. The complete deduced amino acid sequence of ERCC5 was reconstructed from several cDNA clones encoding a predicted protein of 1,186 amino acids. The ERCC5 protein has extensive sequence similarities, in bipartite domains A and B, to products of RAD repair genes of two yeasts, Saccharomyces cerevisiae RAD2 and Schizosaccharomyces pombe rad13. Sequence, structural, and functional data taken together indicate that ERCC5 and its relatives are probable functional homologs. A second locus represented by S. cerevisiae YKL510 and S. pombe rad2 genes is structurally distinct from the ERCC5 locus but retains vestigial A and B domain similarities. Our analyses suggest that ERCC5 is a nuclear-localized protein with one or more highly conserved helix-loop-helix segments within domains A and B.