Chenguang Feng

Positive Darwinian selection within interferon regulatory factor genes of Gymnocypris przewalskii (Cyprinidae) on the Tibetan Plateau

Tibetan Plateau (TP) had experienced phased uplift, resulting in inhospitable environment of low temperature, hypoxia and high ultraviolet radiation for Tibetan wildlife. Many organisms can well adapt to TP, it is of ecological and evolutionary interest to untangle how organisms adapt to extreme environment on TP through evolution. Previous studies mainly focused on hypoxia and metabolism related genes, but we know little about the evolutionary history of immune genes in Tibetan wildlife. In this study, we first identified 10 interferon regulatory factor (IRF) genes from Tibetan naked carp Gymnocypris przewalskii. Within this gene family, IRF3, IRF5, IRF7 and IRF8 contained positive selection sites. Evidences indicated that positive selection may lead to IRF genes functional alternations, presumably driving genes towards adaptation to the environmental changes. Taken together, our results suggested 4 candidate genes as interesting targets for further experimental confirmation of their functional variations and contributions to high altitude adaptation in Tibet fish.

Characterization of two paralogous myostatin genes and evidence for positive selection in Tibet fish: Gymnocypris przewalskii

Myostatin (mstn) is an important member of TGF-β superfamily, a muscle growth inhibitor. Though mstn has been identified in many organisms, little is known about this gene in highland fish, Gymnocypris przewalskii endemic to the Qinghai-Tibetan Plateau. In this study, we first cloned two paralogous mstn genes (mstn1 and mstn2) from G. przewalskii through homologue cloning. The 3D structures of both Mstn proteins varied in the numbers of β-sheets and conformations of α-helices. The branch-site model showed that mstn1 has undergone positive selection, and two positively selected sites (107M and 181T) were located on the random coils of the 3D protein structure. Expression patterns indicated that the mstn1 expressed widely, while the mstn2 only expressed in the muscle and brain. During the early stage of embryo development, the expression levels of both mstn paralogous genes showed different increasing trends. These results suggest that it is diverging in two mstn paralogues of G. przewalskii via specific differences in gene structure, protein structure, selection pressure and gene expression patterns. Taken together, this study provides novel contribution on the research topics of growth related gene function and mechanism of highland fish in extreme aquatic environment on the Qinghai-Tibetan Plateau.

Transcriptomic profiling of Tibetan highland fish (Gymnocypris przewalskii) in response to the infection of parasite ciliate Ichthyophthirius multifiliis

ABSTRACT Gymnocypris przewalskii is a native cyprinid in the Lake Qinghai of the Qinghai‐Tibetan Plateau. G. przewalskii is highly susceptible to the infection of a parasite, Ichthyophthirius multifiliis, in the artificial propagation and breeding. To better understand the host immune reaction to I. multifiliis infection, we characterize the gene expression profiles in the spleen of healthy and I. multifiliis infected G. przewalskii by RNA‐seq. Totally, the transcriptomic analysis produces 463,031,110 high quality reads, which are assembled to 213,538 genes with N50 of 1918 bp and the average length of 1205 bp. Of assembled genes, 90.52% are annotated by public databases. The expression analysis shows 744 genes are significantly changed by the infection of I. multifiliis, which are validated by qRT‐PCR with the correlation coefficient of 0.896. The differentially expressed genes are classified into 689 GO terms and 230 KEGG pathways, highlighting the promoted innate immunity in I. multifiliis infected G. przewalskii at 2 days post infection. Our results pinpoint that the up‐regulated genes are enriched in TLR signaling pathway, inflammatory response and activation of immune cell migration. On the contrary, complement genes are down‐regulated, indicating the evasion of host complement cascades by I. multifiliis. The repressed genes are also enriched in the pathways related to metabolism and endocrine, suggesting the metabolic disturbance in I. multifiliis treated G. przewalskii. In summary, the present study profiles the gene expression signature of G. przewalskii in the responses to I. multifiliis infection, and improves our understanding on molecular mechanisms of host‐parasite interaction in G. przewalskii, which focuses the crucial function of TLRs, cytokines and complement components in the host defense against I. multifiliis. HighlightWe evaluated transcriptomic alterations in G. przewalskii induced by I. multifiliis infection.Totally, 463,031,110 high quality reads were produced by RNA‐seq, which were finally assembled to 213,538 genes.After I. multifiliis challenge, 744 genes were significantly differentially expressed in the spleen of G. przewalskii.The DEGs were enriched in the TLR signaling pathway, complement cascade, and inflammation.The parasite might develop the strategy to repress genes in complement cascades in G. przewalskii.

Taxonomic status of a population of Gymoncypris waddelli Regan, 1905 (Cypriniformes: Schizothoracinae) distributed in Pengqu River, Tibet, China

Gymnocypris waddelli, a highland cold-water fish, is distributed among rivers and lakes of the southern Qinghai-Tibet Plateau. Although previously described as G. waddelli, specimens from the Pengqu River are morphologically distinguishable from those from Lake Yamzhog Yumco, which is the type locality of G. waddelli. We investigated morphological variations of G. waddelli specimens from Pengqu River and Lake Yamzhog Yumco using not only traditional morphological methods but also newly-developed geometric morphometric analysis; furthermore, mitochondrial Cytochrome b (Cyt b) gene analysis was employed to explore the phylogenetic position of the Pengqu River population. Our morphological analyses suggested that G. waddelli from the Pengqu River had an obtuse snout, nearly straight oral fissure, shorter lower jaw than upper jaw, broad mouth and sparse inside gill rakers of the first arch relative to the Lake Yamzhog Yumco population. The strongly supported monophyly of Pengqu population according to mitochondrial datasets was obtained, indicating genetic differences between Pengqu River and Lake Yamzhog Yumco populations. Given their substantial genetic and morphological divergence, specimens from Pengqu River warrant recognition as a distinct species, here described and named Gymnocypris pengquensis.

Comparative cytogenetics of two sympatric species (Gymnocypris, Schizothoracinae) from the Lake Langcuo, Qinghai-Tibet Plateau

Abstract Schizothoracinae is a subfamily of naked carp which is widespread throughout the Qinghai-Tibet Plateau region. Cytogenetic analyses were performed on two sympatric species of naked carp, Gymnocypris chui and G. scleracanthus from the Lake Langcuo, Qinghai-Tibet Plateau. The results showed G. chui and G. scleracanthus had the same chromosome number (2n = 92), but different karyotypes:, (36m + 16sm + 12st + 28a) for G. chui and (26m + 12sm + 18st + 36a) for G. scleracanthus, respectively. The six loci of 45S ribosomal genes (rDNA) were detected through fluorescent in situ hybridization, and their locations were highly conserved between two species. This study depicted the chromosomal characteristics of the Schizothoracinae species and improved our understanding about the relationships and the evolution of two sympatric species.

Adaptive Evolution of the Eda Gene and Scales Loss in Schizothoracine Fishes in Response to Uplift of the Tibetan Plateau

Schizothoracine is the predominant wild fish subfamily of the Tibetan plateau (TP). Their scales, pharyngeal teeth and barbels have gradually regressed with increasing altitude. Schizothoracine have been divided into three groups: primitive, specialized and highly specialized. Ectodysplasin-A (Eda) has been considered as a major gene that contributes to the development of skin appendages. The present study cloned the Eda genes of 51 Schizothoracine fish species which represent the three groups and five Barbinae species. Phylogenetic analyses indicated that Eda may have acted as the genetic trigger for scale loss in the Schizothoracine. Furthermore, 14 single nucleotide polymorphisms (SNPs) and two deletions (18 bp and 6 bp in size), were also detected in the Eda coding sequence of the highly specialized group compared to the primitive group. The same SNPs and two indels result in four non-synonymous and two G-X-Y and 1 XY motif indels, which possibly contribute to significant structure changes in the Eda gene. The domain including (G-X-Y)n motif in the Eda gene is relatively conserved amongst teleosts. Based on the above results, we hypothesize that the evolution of Eda gene might be associated with the scale loss in Schizothoracine fishes in response to the phased uplift of the TP.

Phylogeography of Schizopygopsis stoliczkai (Cyprinidae) in Northwest Tibetan Plateau area

Abstract Schizopygopsis stoliczkai (Cyprinidae, subfamily Schizothoracinae) is one of the major freshwater fishes endemic to the northwestern margin of the Tibetan Plateau. In the current study, we used mitochondrial DNA markers cytochrome b (Cyt b) and 16S rRNA (16S), as well as the nuclear marker, the second intron of the nuclear beta‐actin gene (Act2), to uncover the phylogeography of S. stoliczkai. In total, we obtained 74 haplotypes from 403 mitochondrial concatenated sequences. The mtDNA markers depict the phylogenetic structures of S. stoliczkai, which consist of clade North and clade South. The split time of the two clades is dated back to 4.27 Mya (95% HPD = 1.96–8.20 Mya). The estimated split time is earlier than the beginning of the ice age of Pleistocene (2.60 Mya), suggesting that the northwestern area of the Tibetan Plateau probably contain at least two glacial refugia for S. stoliczkai. SAMOVA supports the formation of four groups: (i) the Karakash River group; (ii) The Lake Pangong group; (iii) the Shiquan River group; (iv) the Southern Basin group. Clade North included Karakash River, Lake Pangong, and Shiquan River groups, while seven populations of clade South share the haplotypes. Genetic diversity, star‐like network, BSP analysis, as well as negative neutrality tests indicate recent expansions events of S. stoliczkai. Conclusively, our results illustrate the phylogeography of S. stoliczkai, implying the Shiquan River is presumably the main refuge for S. stoliczkai.

Elevational diversity gradients of Tibetan loaches: The relative roles of ecological and evolutionary processes

Abstract It is widely believed that species richness patterns (SRPs) are shaped by both ecological and evolutionary processes. However, the relative roles of these processes remain unclear, especially for aquatic organisms. In this study, we integrated ecological and evolutionary measures to tease apart the relative influences of these factors on the SRP of Tibetan loaches along an extensive elevational gradient. We found that the Tibetan loaches displayed a richness pattern that peaked at midelevations. The mean annual temperature (MAT), mid‐domain effect (MDE), and summed age of colonization (SAC, complex of colonization age and colonization frequency) were the main drivers, accounting for 85%, 51%, and 88% of the variations in the SRP, respectively. The three predictors had very high combined effects (MAT‐MDE‐SAC, MAT‐SAC, and MDE‐SAC were 44%, 38%, and 6%, respectively). Our analyses suggested that energy input, time‐for‐speciation, and species dispersal may directly guide the SRP or mediate it by geometric constraints. Conclusively, the SRP of the Tibetan loaches with elevation is the outcome of interactions between biogeographical processes and regional ecological conditions.