| Small heat shock proteins(sHSPs)are diverse and have a lot of members.As molecular chaperones,they participate in protecting organisms and cells from extreme temperatures,hypoxia,ultraviolet irradiation,hunger,heavy metal and so on.sHSPs not only are essential in the organisms to adapt to the environmental stress,but also play a key role in various biological processes.Current study on insect sHSPs mainly confined to the function of a few insects or a few genes.However,a comprehensive system research on the function of sHSP family in insect growth,development and in response to environmental stress is little,so we studied the function of shsp family in the new model:Tribolium castaneum.Real-time quantitative PCR showed that Tcshsps expressed in all developmental stages in Tribolium castaneum.Among them,Tchsp13.2,Tchsp16.0,Tchsp18.3 and Tchsp20.7b highly expressed at early pupa stage.The study found that 10 Tcshsps were all up-regulated in response to heat stress(45?)but not to cold stress(4?).We classified Tcshsps into three groups by sensitive degree of heat stress:strong expression of stress(Tchsp16.0,Tchspl8.3,Tchsp20.7a,Tchsp20.7b,Tchsp20.8 and Tchsp22.2),weak expression of stress(Tchsp13.2)and the no active type(Tchspl9.7,Tchsp21.8 and Tchsp23.6).Probably the strong expression type of shsps may be more engaged in the thermal resistance of insects so as to ensure the body to keep steady in the hot environment.Previously,no active type was called constitutive expression of sHSPs,it could be more involved in maintaining the body basic metabolic or developmental function.Systemic RNAi experiments showed knocking down Tchsp18.3 caused approximately 86.1%insects not to perform the normal metamorphosis from pupa to adult.These insects were teratogenic and died in the pupal stage.While the other 9 Tcshsps after RNAi had no obvious influence on phenotype.However,RNAi of Tchspl9.7 and Tchsp23.6 had 32.8%and 40.6%mortality in Tribolium castaneum respectively.RNAi analysis found 7 Tcshsps made the adult reproduction reduced and 6 Tcshsps decreased the hatching rate of eggs.Among them,knocking down Tchspl8.3 had the most significant effect on adult fecundity.It reduced fertility significantly once Tchsp18.3 was knocked down in females than in males.Further analysis found that Tchspl8.3 exhibited highest expression level in the ovary.After RNAi of Tchsp18.3 showed obvious atrophy of testis and ovary.The results of experiments confirmed that Tchsp18.3 is closely related to maintain the normal function of ovary.In addition to effect on heat stress,these results suggested that shsps also play an important role in the growth and reproduction of Tribolium castaneum.In addition,Tchsp18.3 is also associated with starvation resistance and shorten the adult survival time after RNAi of Tchsp18.3 in starvation treatment.Tchsp18.3 has important function on thermal stress,starvation stress,eclosion and reproduction.So we performed the RNA-sequencing analysis of the transcriptome of hsp18.3 in Tribolium castaneum.Our data showed 569 genes were differentially expressed.Among them,312 genes were up-regulated and 257 genes were down-regulated in the ds-Tchsp18.3 sample.All the detected genes were assigned to 49 GO items.It showed that genes participate in response to stimulus,development and reproduction process are 31,18 and 9,respectively.Some genes associated with these function,such as B lymphocyte-induced maturation protein-1 is involved in the development process,Glucose dehydrogenase participates in reproductive process.Signal pathway analysis found five pathways are greatly influenced,such as glycerophospholipid metabolism pathway,protein digestion and absorption pathway.This research will help us understand more function of insect shsps and lay the foundation for elucidating the signal transduction mechanism of Tchspl8.3. |
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