Investigation On The Physiological Functions Of The LncRNAs Overlapped With Yeast RPL36A And RPL36B Genes

RPL36A and RPL36 B are one of the 59 pairs of paralogous ribosomal proteins in Saccharomyces cerevisiae.Overlapped LncRNA genes are identified in the antisense strand of both RPL36 A and RPL36 B genes,which named AO193 and BO250,respectively.A large portion of AO193 is overlapped with RPL36 A intron,while BO250 is almost completely overlapped with the second exon of RPL36 B gene.Recent studies indicate that antisense LncRNAs play important roles in regulating gene expression.In order to clarify the relationship among AO193,RPL36 A,BO250,and RPL36 B,this study determined the transcriptional and translational levels of these genes in FY251 strain under different physiological conditions.Results demonstrated that the transcriptional levels of sense and anti-sense genes displayed a positively coordinated manner.However,the transcriptional levels are not restrictly correlated to the steady state protein abundance.These preliminary observations hint the possibility that these LncRNAs could be involved in the transcription or post-transcription of ribosomal protein genes,or even at the step of modulating the translatability of some mRNAs.In order to clarify the relationship among RPL36 A,RPL36B,AO193 and BO250,firstly we analyzed the relationship between RPL36 A and RPL36 B.RPL36A and RPL36 B are two copies of paralogous ribosomal proteins.Although there was no effect of RPL36 A on RPL36 B,the transcription level of RPL36 B was significantly higher than that of RPL36 A.Even if the transcription of RPL36 A is increased,the level of RPL36 B transcription will not change significantly.It shows that there are great differences between the functions of RPL36 A and RPL36 B,and they have mutual coordination.The relationship between RPL36 A,RPL36B,AO193 and BO250 transcription levels was detected by quantitative PCR.The results showed that RPL36 A and AO193 were positively related at the transcriptional level.The relationship between RPL36 B and BO250 is more complicated,and the result has not been clarified yet.In order to explore the function of BO250 and AO193,we overexpressed the AO193 and BO250 by a multicopy plasmid and determined the expression of RPL36 A and RPL36 B.The results showed that overexpression of AO193 had no effect on the transcription of RPL36 A.But the transcription of RPL36 B is somewhat promoting.The impact of the overexpression of BO250 on the cell can not appear in normal circumstances.Besides,AO193 and BO250 is not independent,BO250 inhibits the transcription of AO193.Then we reduced the transcription of AO193 and BO250 by modifying the gene structure.The results showed that the transcription of RPL36 A and RPL36 B decreased significantly.The AO193 or BO250 on the multicopy plasmid transferred to those mutant strains.Only RPL36A-flag,RPL36B-ADH1t/p195-BO250's growth can recovered to the wild type.Based on the above results,we hypothesized that the function of BO250 may maintain the normal function of RPL36 A and RPL36 B under stress conditions.From the mRNA level and protein expression of RPL36 A and RPL36 B,we supposed that the effect of BO250 is not base on the transcription and translation level.Instead,it likely that BO250 is a chaperone,by maintaining the correct modification of RNA and shear or maintain the correct modification and folding of the protein and other aspects of the role.Or as a ribosome factor to maintain the normal ribosome biosynthesis.