RNA-binding activity of the rat calmodulin-binding PEP-19 protein and of the long PEP-19 isoform

Abstract

Synthesis of H1˚ histone protein, in the developing rat brain, seems to be regulated mainly at the post-transcriptional level. Since regulation of RNA metabolism depends on a series of RNA-binding proteins, we have been searching for RNA-binding proteins involved in the post-transcriptional regulation of the H1˚ gene. We recently reported isolation, from a cDNA expression library, of an insert encoding a novel protein, the C-terminal half of which is identical to that of PEP-19, a brain-specific protein involved in calcium metabolism. The novel protein was called long PEP-19 isoform (LPI). Herein we show that LPI, as well as PEP-19, can bind H1˚ RNA. Moreover, in order to improve production of functional LPI/PEP-19, we modified the protocol normally adopted for preparing histidine tagged-proteins from bacteria, by adding an additional purification step. We also found that both LPI and PEP can compete for H1˚ RNA binding with PIPPin (CSD-C2), another RNA-binding protein previously discovered in our laboratory. Since PEP19/LPI contain a calmodulin binding domain, we finally investigated whether their ability to bind RNA is affected by calmodulin. Our results show that calmodulin interferes with binding of H1˚ RNA to both PEP-19 and LPI, while it is not able to bind RNA on its own. This finding suggests that calcium/calmodulin may have a role in controlling H1˚ mRNA metabolism in the developing brain.