Disruption of Mitochondrion-To-Nucleus Interaction in Deceased Cloned Piglets

Abstract

Most animals produced by somatic cell nuclear transfer (SCNT) are heteroplasmic for mitochondrial DNA (mtDNA). Oxidative phosphorylation (OXPHOS) in clones therefore requires the coordinated expression of genes encoded by the nuclear DNA and the two sources of mitochondria. Such interaction is rarely studied because most clones are generated using slaughterhouse oocytes of unrecorded origin. Here we traced the maternal lineages of seven diseased and five one-month-old live cloned piglets by sequencing their mtDNA. Additionally by using a 13K oligonucleotide microarray, we compared the expression profiles of nuclear and mtDNA-encoded genes that are involved in mitochondrial functions and regulation between the cloned groups and their age-matched controls (n=5 per group). We found that the oocytes used to generate the cloned piglets were of either the Large White or Duroc background, and oocyte genetic background was not related to the clones’ survival. Expression profiles of mtDNA-encoded genes in clones and controls showed intermixed clustering patterns without treatment or maternal lineage-dependency. In contrast, clones and controls clustered separately for their global and nuclear DNA-encoded mitochondrial genes in the lungs for both the deceased and live groups. Functional annotation of differentially expressed genes encoded by both nuclear and mtDNA revealed abnormal gene expression in the mitochondrial OXPHOS pathway in deceased clones. Among the nine differentially expressed genes of the OXPHOS pathway, seven were down-regulated in deceased clones compared to controls, suggesting deficiencies in mitochondrial functions. Together, these data demonstrate that the coordination of expression of mitochondrial genes encoded by nuclear and mtDNA is disrupted in the lung of diseased clones.

Successful cloning of an adult breeding boar from the novel Chinese Guike No. 1 swine specialized strain

“…Several cloned piglets have been observed with abnormalities, for instance, lacking an anus and tail (Walker et al 2002 ), malformations showed by some important organs (Schmidt et al 2011 ; Park et al 2007 , 2010 ), as well as smaller birth size and lower growth rate (Jiang et al 2007 ). As has been demonstrated so far, such cloned pigs with severe developmental abnormalities displayed considerable dysregulation in the expression of both nuclear DNA- and mitochondrial DNA-inherited genes (Jiang et al 2007 ; Park et al 2007 , 2010 , 2015 ), which may be attributed to the incorrect somatic cell nuclear reprogramming, the variable epigenetic regulation and the deficiencies in the mitochondrial function occurring in a cytoplasm of recipient oocytes (Tian et al 2008 ; Park et al 2015 ). In the present study, all of the six cloned piglets showed normal health at birth and 1 month of age.…”

Does supplementation of oocytes with additional mtDNA influence developmental outcome?

“…Furthermore, by assessing a second generation of offspring that were produced through natural mating, we were again able to use values from natural matings to assess whether any modifications persisted in that generation or whether there was a shift to the norm. A similar strategy was also employed when assessing the first monkeys generated through metaphase II spindle transfer and their subsequent offspring, 45 and to assess DNA methylation and gene expression profiles 46 and nuclear-mtDNA interactions 47 of cloned pigs. This proved a valuable strategy especially when we assessed the biochemical and hematological values in both generations and observed, for example, how the aberrant values for cholesterol were modified in the next generation.…”

The role of mtDNA in oocyte quality and embryo development

“…This would result in the amino acids contributing to each of the 13 mtDNA‐encoded subunits of the electron transfer chain having different configurations, as shown in a study of pig serial nuclear transfer (St John et al, 2005 ). Consequently, when two mitochondrial genomes are present, electron transfer chains can be assembled that have different efficiencies for generating ATP through OXPHOS as demonstrated in a pig model of SCNT where discordant interaction between the nucleus and mtDNA affected OXPHOS gene expression (Park et al, 2015 ); and mouse cybrid (cells derived from the fusion of an mtDNA depleted cell with an enucleated cell) (McKenzie et al, 2003 ) and pig cybrid (Yu et al, 2015 ) models.…”