| Spiders are one of the world's most species-rich animal lineages,and their visual systems are likewise highly diverse.However,our understanding of the genetic basis of these visual systems remains shallow.With 108 described species worldwide,the genus Leptonetela Kratochví,1978 is the largest genus of the family Leptonetidae,collected from China,Vietnam,Georgia,Azerbaijan,Turkey and Greece.They are small haplogyne spiders,characterized by a distinctive 6-eyed pattern with the PME separated from the strongly recurved ALE and PLE.The described spicies are almost known from caves,and the eyes could reduce to four,two or even none.Cave animals have evolved an unusual suite of troglomorphic traits,including the reduction or loss of eyes and pigmentation.Eye reduction and vision loss is the most fascinating trait to scientists and the general public,understanding the genetic basis of which has challenged evolutionary biologists for many years.Hence,the genetic basis of the visual systems in Leptonetela is significant and subservience for us to explore the visual systems of spiders and the mechanisms underlying eye regression in cave-dwelling organisms.Guizhou Province,located in southwestern China,is famous for typical karst landscapes with well-developed and widespread karst caves.The caves have abundant biotic content,such as the most abundant Leptonetela spider species all over the world.L.reticulopecta Lin &Li,2010,L.tetracantha Lin & Li,2010 and a undescribed spider L.sp1 were collected from five caves,respectively,of Suiyang County,Fuquan City and Guiding County,Guizhou.The L.reticulopecta populations with white eyes inhabit the entrance in Mahuang Cave and Shanwang Cave,the eyes of population reduce into six white eyespots in deep zone of Manwang cave.The L.tetracantha populations was disovered in deep Yangzi Cave and Wengshui Dadong Cave zones,and PME absent,ALE and PLE reduced into small white eyespots,contiguous in the population of Yangzi Cave,the eyes absolutely absent in that of Wengshuidadong Cave.And a new species L.sp1 with six eyes was collected in the entrance of Manwang Cave.These materials with multiple eye phenotypes are good subjects for our research.This research is aimed at as followes:(1)Using RNA-seq technology to explore the genetic basis of the Leptonetela;(2)Recovering the genes associated with vision in Leptonetela to know the expression of these genes in cave spiders;(3)Understanding the genetic difference undering eye reduction in spider to lay a foundation for further research on degradation mechanism.The main studies are two folds:(1)Performing bioinformatic analysis and preliminary studying of the eye degeneration mechanism base on the RNA-seq data of Leptonetela;(2)Identifying genes associated with vision via RNA-seq.The main results of this study are as follows:(1)L.sp1 and L.reticulopecta,collected from Manwang Cave and L.tetracantha,collected from Wengshuida Cave have different eye phenotypes.Our study performed RNA-seq sequencing for mixed samples of multiple individual prosomas from the three Leptonetela species using Illumina Hiseq platform.L.sp1,L.reticulopecta and L.tetracantha devo assembled into 27564,32350 and 35454 Unigenes,and 16573,19275,21488 Unigenes got annotation from NR,S-wiss?Prot,GO,COG,KOG,egg NOG,KEGG and Pfam databases.Comparative analysis found that the overall expression patterns of the genes were similar,which supported the morphological relationship among the three species.(2)The RNA-seq data of two samples(L.sp1,with six eyes,and L.reticulopecta,with six white eyespots)from Man Wang cave were merged assembly into 43642 Unigenes,23089 Unigenes were functional annotated,espectively.We have identified 1408 differently expresse d genes(DEGs)between two samples,with 895 up-regulated and 515 down-regulated in eye-reduction species.And 678 genes got annotation.198 DEGs were annotated to GO database,the most enriched are catalytic activity(GO:0003824)(n=88),binding(GO:0005488)(n=72),metabolic process(GO:0008150)(n=83)and cellular process(GO:0008152)(n=69).179 DEGs were annotated to COG database,the top 3 enriched were General function prediction only(n=56),Energy production and conversion(n=22)and amino acid transport and metabolism(n=19).235 DEGs were annotated to KEGG pathway,the top 10 were Oxidative phosphorylation,Retinol metabolism,ECM-receptor interaction,Drug metabolism-cytochrome P450.The enrichent of structural constituent of cuticle(n=69)might be related to integuments of cave spiders,but there was no difference in circadian rhythm,and enrichment analysis showed a significant differences in metabolism between the two spiders.The expression of eight light-interacting genes is significantly different between two samples,including glass,ninac,Omega-crystallin,Kynurenine formamidase,dihydropteridine reductase,Protein Kinase C,RNA-binding protein lark,6-pyruvoyl tetrahydrobiopterin synthase,which may be related to the difference in visual system between the two,but it may only be the overall expression of the prosomas.(3)We screened five types of vision-related genes,including Photoreceptor Specification,Retinal Determination Network,Crystallins,Retinoid Pathway,Invertebrate and Phototran Sduction,from transcriptomes of single individual prosoma samples of L.reticulopecta and L.tetracantha populations with two different eye phenotypes.Using transcriptomic data from eight samples,we identified the expression of all gene families of four types of genes and just didn't detect eya in Retinal Determination Network,suggesting that canonical vision-related genes may also be expressed in Leptonetela spider eyes.There was no significant difference in the expression of most genes in the samples with degraded eyes,suggesting that the dark environment did not significantly affect the expression of most vision-related genes,suggesting that Leptonetela spiders with degraded eyes may still have partial visual function.The paper only contains the preliminary results of the study,and the larboratory is carrying out subsequent data analysis. |
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