Mammalian Polycistronic mRNAs and Disease

Abstract

Our understanding of gene expression has come far since the 'one-gene one-polypeptide' hypothesis proposed by Beadle and Tatum. In this review, we address the gradual recognition that a growing number of polycistronic genes, originally discovered in viruses, are being identified within the mammalian genome, and that these may provide new insights into disease mechanisms and treatment. We carried out a systematic literature review identifying 13 mammalian genes for which there is evidence for polycistronic expression via translation through an internal ribosome entry site (IRES). Although the canonical mechanism of translation initiation has been studied extensively, here we highlight a process of noncanonical translation, IRES-mediated translation, that is a growing source for understanding complex inheritance, the elucidation of disease mechanisms, and the discovery of novel therapeutic targets. Identification of additional polycistronic genes may provide new insights into disease therapy and allow for new discoveries of both translational and disease mechanisms.

Keywords: IRES; ITAF; bicistronic; cap-independent; eIF; polycistronic.

Figure 1. Functional Organization of Bicistronic Genes

A. Two subunits of a multi-subunit complex whose expression is coordinated in a single transcript: each Open Reading Frame (ORF) codes for a specific subunit of a larger protein complex. B. Functionally similar gene products that are differentially co-expressed: a primary protein is expressed through canonical cap-dependent translation while a secondary and functionally similar protein is differentially expressed through a cap-independent mechanism. C. Functionally distinct gene products that have programmatically-related expression: expression of two differentially functioning proteins is coupled with their operation in a particular biological pathway. Category 3 appears with both tandem and overlapping reading frames. D. Signaling proteins generated by stimulus-coupled protease cleavage or by cap-independent translation: two overlapping ORFs code for necessary products for signal transduction, the primary product is a receptor that initiates signal transduction upon ligand binding while the secondary product is a constitutively active signal.

Figure 2. Key Methodology for Identifying IRES-mediated Translation

Confirmation of IRES activity is done by either: 1) insertion of the putative IRES sequence into a dual reporter vector; or 2) in vitro transcription and translation with excess cap analog. Typical steps for exclusion of cryptic promoter activity or alternative splicing: 1) promoterless vector assay 2) in vitro transcription of a single mRNA, and subsequent in vitro translation or cellular transfection 3) northern blotting and reverse transcription PCR spanning the exons.

Figure 3. A Brief Overview of Cap-Dependent Translation and IRES-mediated, Cap-Independent Translation

Mammalian cap-dependent translation functions through the use of all the canonical initiation factors as well as circularization of the mRNA through the interaction of PABP and eIF4G. Key initiation factors are pictured here. Cap-independent IRES-mediated translation is thought to require a subset of ITAFs, an unknown number canonical initiation factors, and may require circularization.