NEW YORK (GenomeWeb News) – In a paper appearing online this month in the journal Biology of Reproduction, researchers from North Carolina State University and the US Department of Agriculture used a combination of gene expression and sequencing approaches to catalog imprinted genes in developing swine fetuses.
The work represents "the most comprehensive survey of imprinted genes in swine to date," according to senior author Jorge Piedrahita, a genomics researcher at NC State, and his co-authors.
Imprinting refers to a situation in which just one allele is expressed for a given gene. That expression is parent-specific, since it comes from either the paternal or maternal copy of the gene. This process is epigenetically regulated by chromatin modifications such as DNA methylation, which keep the non-expressed copy of the gene silent. For instance, in placental animals imprinting is thought to help to strike a compromise between offspring development and maternal health.
"The theory is that paternally expressed genes extract resources from the mother in favor of the fetus, and that maternally expressed genes protect the mother from having too many resources taken away," Piedrahita said in a statement. He noted that as long as these genes are balanced, animals are healthy. But when one set of genes goes awry, offspring may suffer growth and health problems.
Piedrahita and his team decided to explore if, and how, imprinting affected reproduction in pigs by comparing normal, biparental pig fetuses with cloned, uniparental pig fetuses generated by parthenogenesis.
"In order to increase our understanding of the role of imprinted genes in porcine reproductive biology, and to understand how different mammalian species are regulated by imprinting, it is important that a comprehensive analysis of imprinted genes be carried out in swine," they noted. "While there have been several reports of imprinted genes in swine there is still a considerable amount of information missing."
The researchers found that both the cloned fetus and placenta were smaller than normal controls after 28 days of gestation, although they did not detect any obvious morphological differences. In contrast, after 30 days of gestation, they saw obvious differences in the placental features in the parthenote pig fetuses.
The researchers noted that the diminished size of the parthenote fetuses and placentas — a form of intrauterine growth restriction or IUGR — is consistent with the parental conflict hypothesis. Based on their findings, they speculated that paternally expressed genes may become more important for fetal and placental development at late rather than early stages of development.
Next, the researchers used Affymetrix Porcine GeneChip microarrays and/or semi-quantitative PCR to assess gene expression in brain, muscle, liver, and placental tissue from 30 parthenogenetic fetuses that were 30 days old. The microarray work for the study was done by the North Carolina-based commercial RNA profiling and DNA genotyping company Expression Analysis.
By comparing gene expression in the biparental tissues with expression in the clone, parthenote tissues, the researchers pinpointed 25 potential imprinted genes that were detected in at least one of the tissue types tested.
The team also verified imprinting at several candidate genes and looked at mono- and biallelic gene expression in hybrid crosses using an approach called quantitative allelic pyrosequencing to assess fetal brain, carcass, liver, and placental tissue.
Although there are many remaining questions about the function of the putative imprinted genes, the investigators hope the research will serve as a jumping off point for more comprehensive studies of imprinting in pigs.
"[T]here is a disappointing lack of information on the role of these genes in swine fetal and placental development and function," the authors concluded. "It is our hope that this work will stimulate and assist in providing a framework upon research in this critical area that can be expanded."
In particular, Piedrahita noted that the team is interested in exploring why the response to cloning varies for different animal species. "Hopefully if we can discover the ways in which these genes help regulate the flow of resources to the fetus, we can find a way to correct problems when they occur and eliminate IUGR in clones," he said in a statement.