Abstract
RNA-protein (RNP) granules are dynamic membrane-less organelles that form during normal
growth and in response to stress in a reversible manner. One type of RNP granules is
Processing Bodies (P-bodies, or PBs), which contain mRNA together with RNA processing
enzymes. Their roles include sequestration of malfunctioning mRNA destined for degradation
and storing and silencing of mRNAs when not needed. PBs components and behavior are
highly conserved between yeast and human cells. Our understanding of composition and
assembly by liquid-phase separation normal PBs has progressed in the last decade. In contrast,
our knowledge about involvement of PBs in human disease is scarce as is evidence about the
occurrence of aberrant PBs and clearance of normal or aberrant PBS.
In this project, we propose to study variants in a PBs component associated with a
neurodevelopmental disorder that causes intellectual disability. Currently, there is no cure for
such disorders. We hypothesize that the variants cause accumulation of aberrant PBs that,
unlike normal PBs, are not reversible. The human variants carry missense mutations in residues
identical in the yeast protein. Therefore, we started by modeling the effects of these mutations
on RNA processing and accumulation of PBs in yeast. While normal PBs assemble and
disassemble according to cellular needs, our preliminary evidence in yeast cells points to
accumulation of persistent PBs as the major phenotype of these mutations. We propose to
study the effects of these mutations in yeast and in human tissue culture cells, including
neuronal cell lines. The phenotypes we propose to assess are on the accumulation and
dynamics of persistent PBs, including adverse effects they might have on cell function. In
addition, we will explore possible pathways that can clear normal and persistent PBs, such as
macro- and micro-autophagy and ubiquitin-associated degradation. For these studies we will
use a combination of molecular genetics and cellular approaches. Most approaches for studying
RNP granules, granule clearance pathways in yeast and human cells are established in our lab.
Achieving the goals of this proposal would provide novel paradigms on existence and behavior
of persistent PBs and the role they play in a neurodevelopmental disorder. Moreover, identifying
pathways that can clear normal and aberrant PBs would provide novel therapeutic strategies for
a neurodevelopmental disorder associated with intellectual disability, in line with the missions of
the National Institute of Neurological Disorders and Stroke.