The steady state level of MRPL12 in the subject's fibroblasts was reduced to 30% of control value (Fig. 4A and B). The mt-LSU protein ICT1 was also decreased (~ 30% of control values) as was MRPL3 (by 37%) suggesting that a consequence of the MRPL12 mutation is a global defect in assembly of the large ribosomal subunit (mt-LSU). In order to estimate the effect of the MRPL12 mutation on assembly of the whole ribosome, we also tested three proteins of the small ribosomal subunit (SSU), MRPS18B, MRPS25 and DAP3. These were modestly decreased with levels of ~ 60–80% of control (Fig. 4A and B). Correspondingly, 16S and 12S rRNA levels were decreased by 35% and 22% respectively (Fig. 7A). Since porin indicated that there was no compensatory mitochondrial biogenesis and staining of the mitochondrial network with tetramethylrhodamine methyl ester showed no significant alteration in amount or distribution of mitochondria (AR and ZCL unpublished observation) we conclude that the MRPL12 mutation destabilizes the protein resulting in less mt-LSU and to a lesser extent of the small subunit.
control value
steady state level of MRPL12 in the subject's fibroblasts
30%
%
control values
mt-LSU protein ICT1
~ 30%
%
control values
MRPL3
37%
%
proteins of the small ribosomal subunit (SSU)
three
control
MRPS18B, MRPS25 and DAP3
~ 60–80%
%
control
16S
35%
%
control
12S rRNA levels
22%
%