Eric Jan

The regulation of gene expression at the translational level is fundamental for normal cellular homeostasis and survival. Because control of translation is relatively quick and efficient, cells can regulate gene expression quickly in response to environmental stresses. During some cellular stresses such as viral infection, endoplasmic stress (ER) stress and apoptosis, the cell adapts by inhibiting overall protein synthesis. However, it is apparent that a subset of mRNAs is translated under these conditions. These mRNAs, in general, encode for proteins that are important for the cell to adapt to the environmental stress. The identification of these specialized mRNAs is crucial in our understanding of fundamental gene expression mechanisms and how cells respond to stress and viral infection. The primary areas of the lab are to:

1. Examine the mechanisms by which overall protein synthesis is shutoff in response to cellular stress. ER stress and hypoxia are common stresses that occur during the progression of diseases such as cancer and diabetes. By pinpointing the translational control mechanisms underlying these stresses, we will get a better understanding of how cells respond to stress and how these mechanisms may go awry during disease progression.
2. Identify mRNAs that are selectively translated during conditions when overall protein synthesis is shutoff and to elucidate how these specialized mRNAs can bypass the translational control block. Elucidating the translational control mechanisms of these specialized mRNAs is not only important for our overall understanding of gene expression during stress, but may also reveal novel targets for developing therapies against viral infections, stress and cancer.

One of the major interests in the lab is to understand how viruses interact with the host cell. Specifically, we are interested in how viruses hijack the host translational machinery for proper expression of viral proteins at the expense of host gene expression. A current focus is on an unusual non-canonical mechanism of translation found in an internal ribosome entry site (IRES) of the insect virus, the Cricket Paralysis Virus (CrPV) (see Figure below). IRESs are typically long, structured RNA elements, which can directly recruit ribosomes and normally requires a subset of translation initiation factors. Remarkably, unlike translation of the majority of mRNAs, the CrPV IRES mimics a tRNA to initiate translation independent of an AUG start codon, initiator Met-tRNA, and initiation factors. Thus, the IRES evolved specific RNA elements that recruit and hijack the ribosome. Currently, we are using various biochemical approaches to identify and delineate the RNA elements that manipulate specific functions of the ribosome. The study of the CrPV IRES also serves as a model for understanding other IRESs found in some viruses such as hepatitis C virus, HIV and poliovirus and in some cellular IRES such as the oncogene, c-myc, and the angiogenesis factor, VEGF. The secondary structure of the CrPV IRES.

1. Conserved nucleotide positions are shown in uppercase, and nonconserved nucleotides are in lowercase. Numbering refers the nucleotide position within the CrPV RNA genome. Helical regions are indicated by a black dash between nucleotides. The CCU triplet and the GCU alanine codon, which encodes for the first amino acid, occupy the P- and A-sites of the ribosome, respectively. Underlined nucleotides represent the two amino acid residues in the viral capsid protein. PK denotes a pseudoknot structure and SL denotes a stem-loop.
2. Viruses have evolved numerous strategies to efficiently and preferentially amplify their genome in the host cell. The virus has to evade the host antiviral response while at the same time competes for specific host cellular machinery (i.e. the ribosome). By revealing the mechanisms of the battle between the virus and host, we will have a better understanding of the host response to viral infection and may also uncover novel targets for antiviral therapies. Our lab is using the cricket paralysis virus infection as a model system to reveal fundamental virus host interactions. Using a Drosophila as a model host system, we are presently employing microarray-based approaches to identify the host antiviral response. Furthermore, we are also examining the mechanism by which host protein synthesis is shutoff during CrPV infection in order to understand how the virus recruits ribosomes from host mRNAs.
3. Because the CrPV IRES has the unusual ability to recruit ribosomes directly without the aid of initiation factors, we have exploited this property to reconstitute eukaryotic translation in vitro. Using only purified ribosomes, elongation factors, and aminoacyl-tRNAs, we have reconstituted translation in a test tube using a reporter RNA driven by the CrPV IRES. We aim to exploit this approach to explore the regulation of translation elongation by the modification of translation factors (e.g., by phosphorylation) and RNA regulatory elements (e.g., frameshifting). This will allow us to address several important and long-standing questions about the elongation process and its control. The long term goal is to create a framework for elucidating the full repertoire of controls that regulate translation and thus impact biological processes.
4. The endoplasmic reticulum (ER) is a specialized organelle in the cell where proteins destined to be secreted are folded and glycosylated. The balance of folded proteins in the ER has to be tightly regulated for proper cellular function. For instance, the chronic accumulation of unfolded proteins in the ER (called ER stress) is associated with the progression of diabetes and with diseases where specific missense mutations in genes cause the improper folding of proteins in the ER. The cell senses and responds to ER stress by activating a stress response called the unfolded protein response (UPR). One arm of the UPR is a rapid and dramatic inhibition of protein synthesis in order to decrease the accumulation of unfolded proteins in the ER. Paradoxically, the inhibition of protein synthesis triggers the preferential translation of specialized mRNAs, some of which are vital for the cell to adapt to the ER stress. Through microarray-based approaches, we have identified a subset of these mRNAs and a major focus is to elucidate the noncanonical mechanisms by which these mRNAs are translated during ER stress.