Investigation On The Functional Distinction Between The Yeast Paralogous Ribosomal Proteins L36A And L36B As Well As The Biological Significance

Ribosome is generally considered as the factory for protein production playing a important role in cell growth?differentiation and development.The expression of gene is finally through the ribosome.So the ribosome is playing the very important role in the expression of the whole organism.There are a lot of duplicated genes in most eukaryotic genome,Ribosomal protein genes is a typical representative in Saccharomyces cerevisiae.137 ribosome genes code 78 ribosomal proteins,there are.59 pairs copy gene and 19 single copy genes in 137 ribosomal protein genes.This subject is trying to explore the eukaryotes unique ribosomal protein RPL36 two duplicated genes,the RPL36A and RPL36B,duplicate genes to find out whether the gens has special function and meaning in functional and physical.In order to investigate the specific function of ribosome protein between 136A and 136B,we create the yeast with only ribosome protein such as 2A and 2B;Change the sequences of RPL36 gene's promoter,which would interfere the communication between genes,such as ADH1 promoter replaces the promoter of RPL36A,ADH1 replaces the promoter of RPL36B,the promoter of RPL36A replaces the promoter of RPL36B,the promoter of RPL36B replaces the promoter of RPL36A.By spotting assay on the tablet to observe the growth of FY251?2A?2B?ADH1 replace 36A promoter?ADH1 replace 36B promoter,found that only the growth of 2A is slower,the rest is similar with the wild type.Our preliminary experimental results show that the cell with only RPL36A has the growth defects.Ribosome proteins RPL36B in wild type,has the more importance role in the cell.The experimental results of replace the promoter,reveals that Adhl promoter change the expression and function of RPL36A or RPL36B has no significant effect on the growth of cells.Next we continuously cultured cells in liquid medium,detection the growth of cell and the expression of ribosome protein.The experimental results show that in liquid medium,the growth of different genotype strains situation is not as simple as in the solid medium.In the liquid medium,we observed a more varied Detailed changes:the growth trend of 2A is no as good as with the other bacteria;the growth of FY251?2B?ADH1 replace 36A promoter?ADH1 replace 36B promoter,in the early is very similar,but after 30h it showed slighted and repeated different.We suspect that it is due to after 30h later,the accumulation of growth inhibitor and the depletion of easily used carbon source,the cells in order to adapt to the environment,it changes the pathways of energy metabolic,such as glycolysis,alcohol metabolism etc.Because of the influence of the ribosome composition differences,lead to change the gene and path which cope with stress,reflects on different growth rate.At the same time,the results of Western blot shows that at different time points,the expression of ribosome protein RPL36 homologous gene is a dynamic change,suggesting that the moderate ribosome composition and diversity of cell gene expression and adapt to the adverse growing conditions is critical.Then we detect expression of ribosome protein RPL36 homologous gene when at amino acids starvation.Experimental results showed that during amino acids starvation stress,the expression of RPL36A ribosome protein decreased,suggesting that during the amino acid stress,ribosome composited by the protein RPL36 homologous protein and relative abundance are likely to has a certain influence in translating gene which copy with amino acids starvation stress,and this trends is very different from normal medium.The above observations lead us to find a very interesting and meaningful phenomenon,Adhl replace RPL36A promoter will affect the function of the protein.This phenomenon can be seen by the relative abundance of two RPL36 after replacing the promoter of homologous genes in polysome,knock RPL36B gene growth state.And in two different genotype strains,ribosomes assembled by RPL36A protein showed significant differences.This paper through the growth curve?Polysome analysis?western blot?spotting assay?report gene detects the ribosome separation and mass spectrometry analysis,prove homologous ribosomal protein RPL36A and RPL36B has obvious difference function,and this difference is closely associated with its genetic structure,provide a powerful support for ribosome code hypothesis.Through this research,we believe that the homologous ribosomal protein is not simple the function repetited,but every single ribosomal protein has its special function.These results changed the traditional ideas:protein function is determined by the protein amino acid sequence and has opened up a new field for the study of the direction in the protein function.