Decay of rRNA and mRNAs for LH-I and LH-Ⅱ Polypept...
Figure 5 Northern hybridization with probes specific for the LH-Ⅱ，LH-Ⅰ polypeptides，and rRNAs to RNA from a photosynthetic culture of R.capsulatus after treatment with proflavin.Cells were grown photosynthetically on RCV medium for 1 day and then treated with proflavin（100 μg/ml）as described in Materials and Methods.The 0 point represents RNA samples harvested just before addition of proflavin.The points at 5，10，20，40，and 60 min represent samples removed after proflavin treatment.Total RNA was extracted using a mini preparation.Equal amounts of RNA were fractionated on an agarose gel and then transferred to nitrocellulose（LH-Ⅱ）or Gene Screen（LH-Ⅰ，rRNA）and hybridized with 32P-labeled probes for the LH-Ⅱ，LH-Ⅰ，and rRNA as previously described［41］.The probes are listed in Table 1 and Figure 1
One possible mechanism for regulating gene expression in response to oxygen would be to increase the decay rate of certain mRNAs after a shift from anaerobic conditions.To test this hypothesis，we measured the change in the levels of RNA under various conditions.As a control，we measured the half-lives of selected mRNAs after the addition of the transcriptional inhibitor proflavin to the cell culture by using Northern hybridization（see Figure 5）and dot-blot analysis.The genes coding for the LH-Ⅱ proteins had one transcript（0.5 kilobase（kb）），whereas the genes coding for the LH-Ⅰ proteins had two transcripts：a 0.5-kb transcript coding for the LH-Ⅰ proteins only and a 2.6-kb transcript coding for LH-Ⅰ，RC-L，and RC-M proteins and an as yet unidentified open reading frame［39］（see Figure 5）.The half-life of the 0.5-kb LH-Ⅱ mRNA was 24 min，whereas the half-lives of the 0.5-kb LH-Ⅰ mRNA and the 2.6-kb polycistronic mRNA were 19 and 10 min，respectively.These results suggest that，under steady-state photosynthetic growth conditions in the presence of proflavin，the mRNA for the LH-Ⅱ is more stable than the mRNA for the LH-Ⅰ，and that the 0.5-kb mRNA for the LH-Ⅰ proteins alone is more stable than the 2.6-kb mRNA which also codes for the RC-L and RC-M.For the purposes of comparison，we measured the halflife of rRNAs which have the 23S，16S，14S，and SS species（see Figure 5）.Normally the 23S rRNA is unstable and is specifically degraded to the 16S and 14S rRNA during extraction［25］.The half-life of total rRNA was about 24 min，as measured by dot hybridization with pRCl containing 23S，16S，and 5S rRNA as a probe［35］.
We also examined the stability of mRNAs after shifting from photosynthetic to O2 growth.There were no significant changes in the amounts of rRNA after this shift（16S，14S）（see Figure 6）.However，the mRNAs for the LH-Ⅱ and LH-I decreased markedly.The mRNA for LH-Ⅱ decreased with a half-life of 17 min，whereas the mRNA for LH-Ⅰdecreased with a half-life of 23 min.Therefore，the level of mRNA for the LH-Ⅱ is more sensitive to oxygen than is that for the LH-Ⅰ（see Figure 6）.(https://www.daowen.com)
Figure 6 Northern hybridization with DNA probes specific for the LH-Ⅱ and LH-Ⅰpolypeptides and rRNAs to RNA from a photosynthetic culture of R.capsulatus after a shift to O2.Photosynthetic cells were shifted to O2 growth conditions as described in Materials and Methods.RNA extraction and hybridization were as described in the legend to Figure 5
