A disruption of pachytene DNA metabolism in male mice with chromosomally-derived sterility

DNA metabolism was analyzed in spermatocytes of mice that were sterile either because of X-autosome or autosome-autosome translocations, or because of trisomy. In the strains analyzed, spermatogenic development is arrested by metaphase I or soon thereafter. In all such strains, a disruption of the normal pattern of pachytene DNA metabolism occurred. Prepachytene metabolism appeared normal. Disruption was manifest in both the level of endogenously generated nicks during pachytene and in the distribution of nicks among the different DNA sequence classes. Nicking was more intense in the steriles and tended to be randomized in distribution. Satellite DNA underwent pachytene nick-repair in the steriles but not in fertile controls. The repair capacity of spermatocytes from steriles was equal to that of the fertiles; the higher frequency of nicks in the steriles was due to a persistence of nicking activity.     

Aneuploidy induction in mice: construction and use of a tester stock with 100% nondisjunction

A new murine tester stock for primary nondisjunction incorporates three genetically marked Robertsonian translocations with tribrachial homology (TBH): Rb(6.15)1Ald, Rb(4.6)2Bnr, and Rb(4.15)4Rma. The resultant tricentromeric meiotic configuration leads to 100% aneuploid gametes, but the TBH stock can be maintained by intercrossing, through the complementation of nullisomic and disomic gametes. The only neonatal survivors from tescrosses to wild type come from complementation of aneuploid gametes and genetic tests allow wild type gains or losses of Chromosomes 4, 6, and 15 to be distinguished. Alternatively, cytogenetic examination allows products of wild type chromosome gain, with one metacentric, to be separated from chromosome loss with two metacentrics. A pilot study, with 0-2 Gy X-irradiation of oocytes at diakinesis, revealed twelve examples of chromosome loss in wild type gametes but none of chromosome gain and thus provided no evidence for the induction of nondisjunction.     

Identification of the phosphate-translocator from maize mesophyll chloroplasts

Intact maize mesophyll chloroplasts have been isolated in yields of up to 10 mg of chlorophyll per preparation. The chloroplasts were able to reduce 3-phosphoglycerate at a rate of 2.4 mumol of oxygen/min/mg of chlorophyll. This activity was inhibited by preincubating the intact chloroplasts with pyridoxal 5-phosphate. Chloroplast envelopes have been prepared and the protein profile has been obtained on SDS-polyacrylamide gels. The phosphate-translocator from the chloroplast envelope has been identified as a 30kDa polypeptide.     

New mutations at the imprinted Gnas cluster show gene dosage effects of Gsα in postnatal growth and implicate XLαs in bone and fat metabolism but not in suckling

The imprinted Gnas cluster is involved in obesity, energy metabolism, feeding behavior, and viability. Relative contribution of paternally expressed proteins XLαs, XLN1, and ALEX or a double dose of maternally expressed Gsα to phenotype has not been established. In this study, we have generated two new mutants (Ex1A-T-CON and Ex1A-T) at the Gnas cluster. Paternal inheritance of Ex1A-T-CON leads to loss of imprinting of Gsα, resulting in preweaning growth retardation followed by catch-up growth. Paternal inheritance of Ex1A-T leads to loss of imprinting of Gsα and loss of expression of XLαs and XLN1. These mice have severe preweaning growth retardation and incomplete catch-up growth. They are fully viable probably because suckling is unimpaired, unlike mutants in which the expression of all the known paternally expressed Gnasxl proteins (XLαs, XLN1 and ALEX) is compromised. We suggest that loss of ALEX is most likely responsible for the suckling defects previously observed. In adults, paternal inheritance of Ex1A-T results in an increased metabolic rate and reductions in fat mass, leptin, and bone mineral density attributable to loss of XLαs. This is, to our knowledge, the first report describing a role for XLαs in bone metabolism. We propose that XLαs is involved in the regulation of bone and adipocyte metabolism.     

The identification of the site of action of NN′-dicyclohexylcarbodi-imide as a proteolipid in mitochondrial membranes

Mitochondrial membranes were incubated with NN'-dicyclohexyl[(14)C]carbodi-imide, which irreversibly inhibited the partial reactions of oxidative phosphorylation by 95-100%. Solutions of the membranes were analysed on polyacrylamide gels. Of the radioactivity recovered from the gels 90% was shown to be associated with a single protein of molecular weight about 10000. The radioactive protein and associated phospholipid was solubilized from the membrane by extraction with chloroform-methanol mixtures and was concentrated 50-fold by solvent fractionation and adsorption chromatography on Sephadex LH-20. Several protein-radioactivity peaks were obtained by Sephadex LH-20 chromatography. However, 90-100% of the radioactivity in each peak was shown to be associated with a single protein similar to the major radioactive protein observed in electrophoretograms of the membrane solutions. It is concluded that dicyclohexylcarbodi-imide inhibits mitochondrial oxidative phosphorylation by reacting covalently with a group on this chloroform-methanol-soluble protein. The possible role of this protein in oxidative phosphorylation is discussed.