For Creative Research of RNA Biology
Control of gene expression at the RNA level converts to life much of the information stored in our genomes. RNA processing not only determines the fate of all produced proteins, but it also feeds back to regulate gene transcription and post-transcriptional events. We are interested in understanding how control of gene expression at the RNA level allows cells to make decisions, respond to the environment, and communicate with one another, as well as how misregulation of RNA processing leads to cellular dysfunction and disease. We pursue these problems in two major biological settings. First, we study effector roles of RNA in the brain, especially the mechanisms and phenotypic consequences of post-transcriptional RNA processing, ranging from alternative splicing to protein translation. We are particularly interested in the activities of RNA-binding proteins (RBPs), their regulation, and their dynamic formation of ribonucleoprotein complexes during early neuronal development and in neurodegeneration. Most recently, we have developed a platform to study participation of RBPs in inter-neuronal communication controlled by light. Second, we investigate regulatory roles of RNA in stem cells and during cancer development, especially as it pertains to non-coding RNA-dependent control of gene transcription. Our studies make heavy use of genome-wide and computational approaches (including techniques such as iCLIP, ribosome profiling, RNA-seq, ChIP-seq, and interactome capture) in combination with traditional biochemistry and genetics to allow for comprehensive description of RNA networks and their regulating RBPs. We are also integrating in vitro stem cell differentiation, CRISPR-based strategies, microfluidics and optogenetics to build light-controlled platforms for studies of RNA biology in neurological disorders.
Some of our recent stories
A molecular program that determines cell shape
Cell shape is one of the most distinctive features of all somatic cells yet little is known about how it is specified. We discovered a molecular program, controlled at the RNA level, that determines the early shape of neurons. We found that a single molecule, a deeply conserved RNA-binding protein, called Unkempt, coordinates an entire morphology program to establish the early bipolar shape of neurons during embryonic development of an organism. Intriguingly, we found that this program also has the capacity to confer a similar morphology to cells of different lineages.
To understand what Unkempt-regulated processes might contribute to remodeling of cellular architecture, we first used iCLIP to define the identities and precise binding sites of Unkempt on its targeted RNA molecules. We found that Unkempt functions as a sequence-specific RNA-binding protein that targets coding regions of a defined set of ubiquitously expressed messages linked to protein metabolism and regulation of the cytoskeleton. RNA binding is required for Unkempt-induced remodeling of cellular shape and, as revealed by ribosome profiling, is directly coupled to a reduced production of the encoded proteins. Thus, during embryonic development, Unkempt controls a translationally regulated cell morphology program to ensensure proper structuring of the nervous system. These findings link post-transcriptional regulation of gene expression with
cellular shape and have general implications for the development and disease of multicellular organisms. See the paper here.
Our work on the cover page at G&D
Neuronal morphogenesis via a novel mode of RNA sequence recognition
The CCCH zinc finger proteins constitute the second largest group of RNA-binding proteins in mammals, yet their biology remains poorly understood. Following up on our initial description of Unkempt protein in specifying the early neuronal shape , we wondered how the six CCCH zinc fingers of Unkempt - the largest such array of any mammalian protein - operate to recognize a relatively short stretch of RNA sequence. Structural and functional studies revealed the six zinc fingers form two compact RNA-binding folds, each consisting of three zinc fingers, with no similarity to any other structure in the annotated proteomes. We found that each zinc finger cluster functions as a separate RNA-binding unit that recognizes a distinct trinucleotide RNA motif, either UAG or a U-rich motif. Presence of both motifs and their relative position within coding regions of targeted genes is critical for high-affinity RNA binding, which, in turn, is required for translational repression of Unkempt-target genes, as well as for the induction of bipolar neuronal morphology.
Abstraction of neuronal morphogenesis via RNA binding
The evolutionary origins of the compact CCCH zinc finger clusters of Unkempt can be traced back roughly 600 million years to the emergence of metazoans and the evolution of the neuronal lineage. We found that RNA sequence specificity of Unkempt orthologs as well as their capacity to polarize cells have also remained conserved, suggesting an evolutionary link between the CCCH clusters of Unkempt and neuronal morphology. Given the high abundance of CCCH-type RNA-binding proteins and their wide functional diversity in different organisms, it will be important to determine the prevalence of such clusters, the rules that predict their formation and the set of properties beyond RNA binding that they may impart to proteins. We hope that our work would stimulate and provide some guidance to these endeavors.
Protein methylation in regulation of gene expression
Post-translational modifications (PTMs) of proteins regulate almost every aspect of cell biology by altering the functional properties of proteins, commonly in a reversible manner and at a relatively low energetic cost. Protein methylation is widespread and is perhaps one of the most functionally versatile forms of PTMs, with the capacity to impinge on essentially any cellular process. Accordingly, deregulation of protein methylation has been associated with a plethora of human disorders. We reviewed the field of protein methylation, paying particular attention to its established impact on gene transcription via histone methylation, as well as its less understood effects on post-transcriptional processing via methylation of RNA-binding proteins (see the paper here ). Curiously, a substantial number of methylated sites in RNA-binding proteins appear to be buried deep within protein structure where they are unlikely to be accessible to methyl reader proteins. Buried methylated sites are most commonly located in the immediate vicinity of the bound RNA, which suggests that protein methylation could directly affect protein–RNA interactions. However, the significance of these methylation events remains largely unexplored. Our lab is interested in understanding how methylation and phosphorylation of RNA-binding proteins affects their activities in vivo, the kinetics of these PTMs under physiological conditions, and their misregulation in human disorders.