4.1 Adaptive mechanism of hearing related genes
Rhinolophids arguably possess the most sophisticated echolocation systems, and can emit relatively long calls adapted to detect and classify the wing beats of insects. They are heavily reliant on hearing for a variety of ecologically important roles. Previous studies have documented that hearing-related genes are predominantly evolutionarily conserved in mammals (Kirwan et al., 2013). Comparative analysis of auditory perception can help to elucidate the molecular basis that underpins different auditory capabilities (Dong et al., 2016). Dong et al. (2016) revealed some hearing-related genes undergone natural selection associated with the evolution of specialized constant frequency echolocation, probably resulting from the extreme selectivity used in the auditory processing by these bats. We performed comparative transcriptome for species with two echolocation modes, and tested some genes under selective pressure for the species within themacrotis group.
We selected some specific amino acid changes in six PSGs (CRYM ,FOXM1 , MAP6 , PYCARD , SLC35A2, andWRB ) related to hearing. CRYM was tested in R. marshalli , and the previous functional study found mutations inCRYM may cause hearing loss and the expression of CRYM is essential for maintaining cochlear cells and preserving normal hearing (Abe et al., 2003; Hosoya et al., 2016). We testedFOXM1 , MAP6 , and WRB in R. episcopus . Previous studies showed that FOXM1 activities are modulated in the mouse cochlea, and FOXM1 is associated with cell cycle control and essential for the transcriptional response during DNA damage/checkpoint signaling (Sanchez-Calderon et al., 2010).MAP6 is involved in molecular transport, nervous system development, and function, and is related to reflect auditory fear conditioning (Hong et al., 2013). The absence of WRB from inner hair cells results in significantly reduced intracellular levels of otoferlin, thus causing hair cell synaptic disruption and hearing impairment (Pangrsic and Vogl, 2018). PYCARDand SLC52A2 were tested in R. siamensis . Thereinto,PYCARD was related to hearing-loss and tinnitus (Wei et al., 2010). Mutation inSLC52A2 was associated with spinocerebellar ataxia with blindness and deafness type2 (O’Callaghan et al., 2019). We tested different hearing related genes in different specieslow, and this may imply these species adapted to the environment through the different evolutionary mechanisms.
Particularly, we found one PSG SPRY2 in R. osgoodi , who emit echolocation with normal frequency, differ from other species within the macrotis group. SPRY2 is related to hearing loss and plays an important role in the regulation of endochondral bone formation, which may influence early inner ear development (Joo et al., 2016; Wright et al., 2015; Yousaf et al., 2018). The positive selection sites detected in SPRY2 showed similar amino acid changes with R. pusillus , but different with other species. This result indicated that SPRY2 plays an important role in the echolocation development, and may experience similar acoustic evolution in R. pusillus , thus promote R. osgoodi and R. pusillus evolved similar acoustic characteristics.
Although the function of specific amino acid variants in these genes is still unclear, our results still indicate the potentially important function in acoustic development. These genes may regulate the neural activity or the formation process of inner ear structure, and then affect the sensitivity of specific frequency, that is, affect the echolocation signal for these bats.