Identification of genes showing differential expression profile associated with growth rate in skeletal muscle tissue of Landrace weanling pig.

Suppression subtractive hybridization was used to identify genes showing differential expression profile associated with growth rate in skeletal muscle tissue of Landrace weanling pig. Two subtracted cDNA populations were generated from musculus longissimus muscle tissues of selected pigs with extreme expected breeding values at the age of 100 kg. Three upregulated genes (EEF1A2, TSG101 and TTN) and six downregulated genes (ATP5B, ATP5C1, COQ3, HADHA, MYH1 and MYH7) in pig with genetic propensity for higher growth rate were identified by sequence analysis of 12 differentially expressed clones selected by differential screening following the generation of the subtracted cDNA population. Real-time PCR analysis confirmed difference in expression profiles of the identified genes in musculus longissimus muscle tissues between the two Landrace weanling pig groups with divergent genetic propensity for growth rate. Further, differential expression of the identified genes except for the TTN was validated by Western blot analysis. Additionally, the eight genes other than the ATP5C1 colocalized with the same chromosomal positions as QTLs that have been previously identified for growth rate traits. Finally, the changes of expression predicted from gene function suggested association of upregulation of expression of the EEF1A2, TSG101 and TTN genes and downregulation of the ATP5B, ATP5C1, COQ3, HADHA, MYH1 and MYH7 gene expression with increased growth rate. The identified genes will provide an important insight in understanding the molecular mechanism underlying growth rate in Landrace pig breed.

Mechanism on two-electron oxidation of ubiquinol at the Qp site in cytochrome bc1 complex: B3LYP study with broken symmetry.

The molecular and electronic structures of the Rieske iron-sulfur [2Fe-2S] cluster with an imidazolate and imidazole were investigated by using usual unrestricted and broken symmetry B3LYP methods for the highest and lowest spin states, respectively. The electronic structures of the lowest spin states were determined by the spin contaminations and natural orbital analyses. It was shown that the spin contamination presents the number of pairs of the antiferromagnetic spin couplings. The oxidation mechanism of the ubquinol at the Q(p) site of the cytochrome bc(1) complex was also examined by the broken symmetry B3LYP methods. In the [2Fe-2S] clusters with an imidazolate, the oxidized and lowest spin state, [(Imz(-))FeS](ox)LS, was lowest in energy among four possible states, consistent with experimental observations. In the examination of the mechanism of the ubquinol oxidation, it was confirmed that the ubiquinol docks between the imidazolate of [2Fe-2S] clusters and Glu272(-) of cytochrome b by the hydrogen bonds before the oxidation proceeds, consistent with the experimental proposals. Our results support a "Glu272-first" mechanism that Glu272 serves as an acceptor of the first proton from the ubiquinol and subsequently the proton-coupled electron transfer (PCET) occurs from the ubisemiquinone anion to the Rieske iron-sulfur [2Fe-2S] cluster.