Gene expression vs. sequence divergence: comparative transcriptome sequencing among natural Rhinolophus ferrumequinum populations with different acoustic phenotypes

Abstract

Background Although the sensory drive hypothesis can explain the geographic variation in echolocation frequencies of some bat species, the molecular mechanisms underlying this phenomenon are still unclear. The three lineages of greater horseshoe bat (Rhinolophus ferrumequinum) in China (northeast, central-east, and southwest) have significant geographic variation in resting frequencies (RF) of echolocation calls. Because their cochleae have an acoustic fovea that is highly sensitive to a narrow range of frequencies, we reported the transcriptomes of cochleae collected from three genetic lineages of R. ferrumequinum, which is an ideal organism for studying geographic variation in echolocation signals, and tried to understand the mechanisms behind this bat phenomenon by analyzing gene expression and sequence variation. Results A total of 8190 differentially expressed genes (DEGs) were identified. We identified five modules from all DEGs that were significantly related to RF or forearm length (FL). DEGs in the RF-related modules were significantly enriched in the gene categories involved in neural activity, learning, and response to sound. DEGs in the FL-related modules were significantly enriched in the pathways related to muscle and actin functions. Using 21,945 single nucleotide polymorphisms, we identified 18 candidate unigenes associated with hearing, five of which were differentially expressed among the three populations. Additionally, the gene ERBB4, which regulates diverse cellular processes in the inner ear such as cell proliferation and differentiation, was in the largest module. We also found 49 unigenes that were under positive selection from 4105 one-to-one orthologous gene pairs between the three R. ferrumequinum lineages and three other Chiroptera species. Conclusions The variability of gene expression and sequence divergence at the molecular level might provide evidence that can help elucidate the genetic basis of geographic variation in echolocation signals of greater horseshoe bats.

Echolocation call frequency variation in horseshoe bats: molecular basis revealed by comparative transcriptomics

“…Results from these studies and, more recently, genome-wide screens have revealed that multiple hearing genes show molecular adaptation in lineages of echolocators, in some cases involving convergent amino acid replacements [20][21][22][23][24]. Despite insights from such comparative sequencing studies, little is known about the genetic basis of divergence in echolocation call frequencies among populations and closely related species of bats, including the role of gene expression difference (but see [25]).…”

“…[17,18,23,36]). At the intra-specific level, Zhao et al [25] were the first to study the genetic basis of variation in echolocation call frequencies in three geographical populations of the greater horseshoe bat (R. ferrumequinum) using comparative transcriptome sequencing. However, in this study, variation of call frequencies among the focal populations was slight (less than 8 kHz), potentially limiting the sensitivity of such an approach for distinguishing true candidates.…”

Transcriptome sequencing of cochleae from constant-frequency and frequency-modulated echolocating bats

“…Modulation of gene expression and alternative mRNA splicing are two major forms of transcriptional regulation, responsible for the origin of novel phenotype and phenotypic diversity 4 – 7 . Recently, high-throughput transcriptome sequencing (RNA-seq) of cochlear tissue has been used to uncover differentially expressed genes possibly associated with the origin of ultrahigh frequency hearing 8 , the divergence of different echolocating types 9 and echolocation call frequency variation 10 . In these earlier studies, the reference used for quantification of gene expression was from a de novo assembly based on the short RNA-seq reads which may contain many artificial transcripts 11 .…”

“…The current FL transcriptomes generated in this study are sufficient to be reused in the several aspects. They can be used as the reference to reanalyze the RNA-seq datasets of cochlea in previous comparative transcriptomic studies 8 – 10 . Quantification of transcript expression by mapping reads to the FL transcriptome will help to improve the accuracy of identifying differentially expressed transcripts 12 .…”

Assembly-free rapid differential gene expression analysis in non-model organisms using DNA-protein alignment

“…On the other hand, driven by declining costs, RNA-seq is becoming increasingly accessible to labs with modest resources; and as a result, it is being employed on an everexpanding catalog of non-model organisms, pervading the fields of agriculture, aquaculture, ecology, and environment. A very short list of recent studies include: environmental stress response in sea-trout [3], coral [4], ryegrass [5], pigeonpea [6], tiger barb [7]; immune response to parasites and pathogens in guppy [8], eel [9], silkworm [10], peanut [11], sunflower [12]; mechanisms of phenotypic divergence in hares [13], bats [14], grass carps [15]; effect of diet in the growth and development in shrimp [16], yellow perch [17], mandarin fish [18], grenadier anchovy [19], catfish [20], tilapia [21], bass [22]. It is only likely that RNA-seq will continue to rapidly proliferate while high-quality reference databases grow at a slow pace.…”