rRNA inheritance in neurons is sufficient for neurodevelopment and
protein synthesis.
We next asked if inhibition of rRNA synthesis would differentially
affect neural progenitors and neurons. To achieve cell type-specific
inhibition of rRNA synthesis, we used a previously characterized RNA
interference line [7] to knockdown RNA polymerase I subunit B(Polr1B ) in progenitors (insc-Gal4 x UAS-Polr1B{RNAi} )
or neurons (nSyb-Gal4 x UAS- Polr1B{RNAi}) . Polr1Bknockdown using insc-Gal4 limits rRNA inheritance in neurons but
does not affect rRNA synthesis in neurons (Figure 4A ).Polr1B knockdown using nSyb-Gal4 limits rRNA synthesis in
neurons but does not affect rRNA inheritance (Figure 4A ). As
previously described for a RNAi screen in neuroblasts [23], we found
that Polr1B knockdown driven by insc-Gal4 limits
neurogenesis and causes nearly 100% failure of adult flies to eclose
from their pupal case (Figure 4B ). Insc-Gal4 x
UAS-Polr1B{RNAi} flies manually dissected from their pupal case were
alive but unable to walk or fly, indicative of neurologic defects. In
contrast, Polr1B knockdown in neurons had no effect on
development: flies eclosed 100% of the time (Figure 4B ) and
exhibited normal motor function.
Next we tested if inherited rRNA is sufficient to support bulk protein
synthesis in neurons. We used a fluorophore-“clickable” amino acid
analog (homopropargylglycine (HPG)) [31] to quantify protein
synthesis in neurons of wildtype, insc-Gal4 x UAS-
Polr1B{RNAi}, and nSyb-Gal4 x UAS- Polr1B{RNAi} flies.
Dissected brains were soaked in HPG for three hours to allow sufficient
labeling while limiting the likelihood of proteins passing from GMCs to
neurons at cytokinesis (based on the 4.2-hour average GMC cell cycle).
As previously shown [23], Polr1B knockdown usinginsc-Gal4 decreases progenitor proliferation, resulting in
smaller brain size, but neurons are still produced (Figure 4C ).
These neurons, with intact autonomous rRNA synthesis but limited rRNA
inheritance, had significantly reduced levels of protein synthesis
(Figures 4C and D ). Neurons with intact rRNA inheritance but
limited autonomous rRNA synthesis had wildtype levels of protein
synthesis (Figures 4C and D ). We interpret these data as
evidence that inherited rRNAs significantly contribute to protein
synthesis in neurons.