Transcriptome, genetic editing, and microRNA divergence substantiate sympatric speciation of blind mole rat, Spalax.

Incipient sympatric speciation in blind mole rat, Spalax galili, in Israel, caused by sharp ecological divergence of abutting chalk-basalt ecologies, has been proposed previously based on mitochondrial and whole-genome nuclear DNA. Here, we present new evidence, including transcriptome, DNA editing, microRNA, and codon usage, substantiating earlier evidence for adaptive divergence in the abutting chalk and basalt populations. Genetic divergence, based on the previous and new evidence, is ongoing despite restricted gene flow between the two populations. The principal component analysis, neighbor-joining tree, and genetic structure analysis of the transcriptome clearly show the clustered divergent two mole rat populations. Gene-expression level analysis indicates that the population transcriptome divergence is displayed not only by soil divergence but also by sex. Gene ontology enrichment of the differentially expressed genes from the two abutting soil populations highlights reproductive isolation. Alternative splicing variation of the two abutting soil populations displays two distinct splicing patterns. L-shaped FST distribution indicates that the two populations have undergone divergence with gene flow. Transcriptome divergent genes highlight neurogenetics and nutrition characterizing the chalk population, and energetics, metabolism, musculature, and sensory perception characterizing the abutting basalt population. Remarkably, microRNAs also display divergence between the two populations. The GC content is significantly higher in chalk than in basalt, and stress-response genes mostly prefer nonoptimal codons. The multiple lines of evidence of ecological-genomic and genetic divergence highlight that natural selection overrules the gene flow between the two abutting populations, substantiating the sharp ecological chalk-basalt divergence driving sympatric speciation.

Adaptive methylation regulation of p53 pathway in sympatric speciation of blind mole rats, Spalax.

Epigenetic modifications play significant roles in adaptive evolution. The tumor suppressor p53, well known for controlling cell fate and maintaining genomic stability, is much less known as a master gene in environmental adaptation involving methylation modifications. The blind subterranean mole rat Spalax eherenbergi superspecies in Israel consists of four species that speciated peripatrically. Remarkably, the northern Galilee species Spalax galili (2n = 52) underwent adaptive ecological sympatric speciation, caused by the sharply divergent chalk and basalt ecologies. This was demonstrated by mitochondrial and nuclear genomic evidence. Here we show that the expression patterns of the p53 regulatory pathway diversified between the abutting sympatric populations of S. galili in sharply divergent chalk-basalt ecologies. We identified higher methylation on several sites of the p53 promoter in the population living in chalk soil (chalk population). Site mutagenesis showed that methylation on these sites linked to the transcriptional repression of p53 involving Cut-Like Homeobox 1 (Cux1), paired box 4 (Pax 4), Pax 6, and activator protein 1 (AP-1). Diverse expression levels of p53 between the incipiently sympatrically speciating chalk-basalt abutting populations of S. galili selectively affected cell-cycle arrest but not apoptosis. We hypothesize that methylation modification of p53 has adaptively shifted in supervising its target genes during sympatric speciation of S. galili to cope with the contrasting environmental stresses of the abutting divergent chalk-basalt ecologies.

Codon 104 variation of p53 gene provides adaptive apoptotic responses to extreme environments in mammals of the Tibet plateau.

Mutational changes in p53 correlate well with tumorigenesis. Remarkably, however, relatively little is known about the role that p53 variations may play in environmental adaptation. Here we report that codon asparagine-104 (104N) and glutamic acid-104 (104E), respectively, of the p53 gene in the wild zokor (Myospalax baileyi) and root vole (Microtus oeconomus) are adaptively variable, meeting the environmental stresses of the Tibetan plateau. They differ from serine-104 (104S) seen in other rodents, including the lowland subterranean zokor Myospalax cansus, and from serine 106 (106S) in humans. Based on site-directed mutational analysis in human cell lines, the codon 104N variation in M. baileyi is responsible for the adaptive balance of the transactivation of apoptotic genes under hypoxia, cold, and acidic stresses. The 104E p53 variant in Microtus oeconomus suppresses apoptotic gene transactivation and cell apoptosis. Neither 104N nor 104E affects the cell-cycle genes. We propose that these variations in p53 codon 104 are an outcome of environmental adaptation and evolutionary selection that enhance cellular strategies for surviving the environmental stresses of hypoxia and cold (in M. baileyi and M. oeconomus) and hypercapnia (in M. baileyi) in the stressful environments of the Qinghai-Tibet plateau.

Growth hormone and insulin-like growth factor of naked carp (Gymnocypris przewalskii) in Lake Qinghai: expression in different water environments.

Here, we report the cloning and characterization of growth hormone (GH), insulin-like growth factor-I (IGF-I) and IGF-II from naked carp (Gymnocypris przewalskii), a native teleost fish of Lake Qinghai in the Qinghai-Tibet Plateau of China. The GH of naked carp encodes for a predicted amino acid sequence showing identities of 63%, 63%, 91% and 94% with cherry salmon, rainbow trout, zebrafish and grass carp, respectively. Compared to common carp and goldfish, evolutionary analysis showed that genome duplication has had less influence on the relaxation of purifying selection in the evolution of naked carp GH. Sequence analysis of naked carp IGF-I (ncIGF-I) and ncIGF-II showed a high degree of homology with known fish IGF-I and IGF-II. To investigate effects of salinity and ionic composition of the aquatic environment on the GH-IGF axis in naked carp, male fish held in river water were assigned randomly to 4 groups: RW (river-water), RW+Na (NaCl in RW), RW+Mg (MgCl(2) in RW) and LW (lake-water) groups. The concentrations of Na(+) in RW+Na and Mg(2+) in RW+Mg were equal to the concentrations of these ions in lake-water. After 2 days of exposure, the plasma IGF-I levels in the RW+Na and LW groups were significantly higher than the control group (RW), and the plasma GH levels of the LW group were also significantly higher than the RW group. The somatostatin (SS) levels in the hypothalamus significantly increased in the RW+Na group. After 5 days of exposure, these hormone levels did not differ significantly among groups. These results indicate that while the plasma GH and IGF-I levels are osmosensitive, the absence of a change in GH secretion in RW+Na might be partly due to a transiently increased release of hypothalamic SS induced by the stress of neutral-saline water. This is the first report of a salinity-induced increase of GH-IGF-I circulating levels in Cypriniformes.

Evolution and regulation of the downstream gene of hypoxia-inducible factor-1alpha in naked carp (Gymnocypris przewalskii) from Lake Qinghai, China.

The naked carp (Gymnocypris przewalskii) is a native teleost of Lake Qinghai (altitude, 3.2 km) on the Qinghai-Tibet Plateau in China. Hypoxia-inducible factor (HIF)-1alpha of Gymnocypris przewalskii was cloned and a phylogenetic tree for vertebrate HIF-1alpha was constructed. By analysis of maximum likelihood models of codon substitutions for HIF-1alpha, three positive sites in the branch lineages of crucian carp, eelpout, and flounder, and a higher proportion of neutral sites in naked carp, antarctic eelpout, rainbow trout, and grayling, were detected among all teleosts. It seems that low habitat temperatures relax the purifying selection of HIF-1alpha in these stenothermal coldwater fish, and both cold and hypoxic lake water contributed to the evolution of the HIF-1alpha gene in the naked carp. Furthermore, Glut 1 mRNA, a gene downstream from HIF-1alpha, has a time-course- and tissue-specific dependent response to hypoxic challenge.