Protein synthesis is often regulated at the level of initiation of translation, making it a critical step. This regulation occurs by both the cis-regulatory elements, which are located in the 5 -and 3 -UTRs (untranslated regions), and trans-acting factors. A breakdown in this regulation machinery can perturb cellular metabolism, leading to various physiological abnormalities. The highly structured UTRs, along with features such as GC-richness, upstream open reading frames and internal ribosome entry sites, significantly influence the rate of translation of mRNAs. In this review, we discuss how changes in the cis-regulatory sequences of the UTRs, for example, point mutations and truncations, influence expression of specific genes at the level of translation. Such modifications may tilt the physiological balance from healthy to diseased states, resulting in conditions such as hereditary thrombocythaemia, breast cancer, fragile X syndrome, bipolar affective disorder and Alzheimer's disease. This information tends to establish the crucial role of UTRs, perhaps as much as that of coding sequences, in health and disease.

We have cloned the cDNA of the hemeregulated eIF-2a kinase (HRI) of rabbit reticulocytes. In vitro translation of mRNA transcribed from the HRI cDNA yields a 90-kDa polypeptide that exhibits eIF-2a kinase activity and is recognized by a monoclonal antibody directed against authenticHRI. The open reading frame sequence of the HRI cDNA contains all 11 catalytic domains of protein kinases with consensus sequences of protein-serine/threonine kinases in conserved catalytic domains VI and Vm. The HRI cDNA also contains an insert of -140 amino acids between catalytic domains V and VI. The HRI cDNA coding sequence has extensive homology to GCN2 protein kinase of Saccharomyces cerevisiae and to human double-stranded-RNA-dependent eIF-2a kinase. This observation suggests that GCN2 protein kinase may be an eIF-2a kinase in yeast. In addition, HRI has an unusually high degree of homology to three protein kinases (NimA, Weel, and CDC2) that are involved in the regulation of the cell cycle.Protein synthesis in intact reticulocytes and their lysates is dependent on the availability of heme (1-4). In heme deficiency, protein synthesis is inhibited at the level of initiation due to the activation ofa heme-regulated inhibitor (HRI), also called the heme-controlled repressor (refs. 4-6; for review, see refs. 7 and 8). HRI is a cAMP-independent protein kinase that specifically phosphorylates eIF-2a (9-12). Phosphorylation of eIF-2a [eIF-2(aP)] in reticulocyte lysates results in the binding and sequestration of reversing factor (RF), also designated as guanine nucleotide exchange factor or eIF-2B, in an RF-eIF-2(aP) complex. Since RF is required for the exchange of GTP for GDP in the recycling of eIF-2 and in the formation of the eIF-2-Met-tRNAMet.GTP ternary complex, its unavailability results in the cessation of the initiation of protein synthesis (ref. 13; for review, see refs. 7 and 8).Although the mechanism of regulation of protein synthesis by HRI has been extensively studied, little is known about the structure and regulation of HRI itself. We have reported (14) the amino acid sequences of three tryptic peptides of heme-reversible HRI. HRI peptide P-52 contains the sequence Asp-Phe-Gly, which is the most highly conserved short stretch in the conserved domain VII of protein kinases as presented by Hanks et al. (15). The N-terminal 14 amino acids of HRI peptide P-74 show 50-60%o identity to the conserved domain IX of kinase-related transforming proteins (14). These findings are consistent with the autokinase and eIF-2a kinase activities of HRI. To study the structurefunction relations of HRI, we undertook the molecular cloning of rabbit HRI cDNA,l which we report here. METHODSPCR Amplification of HRI cDNA Between P-52 and P-74. Poly(A)+ mRNA (1 ,ug) was reverse-transcribed to obtain single-stranded cDNAs (16). The sense-strand oligodeoxynucleotide of P-52 and the antisense-strand oligodeoxynucleotide of P-74, deduced with preferred codon usage (17), were used as primers. PCR was carried out in the presence of single-st...

How tumour suppressor p53 bifurcates cell cycle arrest and apoptosis and executes these distinct pathways is not clearly understood. We show that BAX and PUMA promoters harbour an identical MAR element and are transcriptional targets of SMAR1. On mild DNA damage, SMAR1 selectively represses BAX and PUMA through binding to the MAR independently of inducing p53 deacetylation through HDAC1. This generates an anti-apoptotic response leading to cell cycle arrest. Importantly, knockdown of SMAR1 induces apoptosis, which is abrogated in the absence of p53. Conversely, apoptotic DNA damage results in increased size and number of promyelocytic leukaemia (PML) nuclear bodies with consequent sequestration of SMAR1. This facilitates p53 acetylation and restricts SMAR1 binding to BAX and PUMA MAR leading to apoptosis. Thus, our study establishes MAR as a damage responsive cis element and SMAR1-PML crosstalk as a switch that modulates the decision between cell cycle arrest and apoptosis in response to DNA damage.

Cellular mRNAs are predominantly translated in a cap-dependent manner. However, some viral and a subset of cellular mRNAs initiate their translation in a cap-independent manner. This requires presence of a structured RNA element, known as, Internal Ribosome Entry Site (IRES) in their 5′ untranslated regions (UTRs). Experimental demonstration of IRES in UTR remains a challenging task. Computational prediction of IRES merely based on sequence and structure conservation is also difficult, particularly for cellular IRES. A web server, IRESPred is developed for prediction of both viral and cellular IRES using Support Vector Machine (SVM). The predictive model was built using 35 features that are based on sequence and structural properties of UTRs and the probabilities of interactions between UTR and small subunit ribosomal proteins (SSRPs). The model was found to have 75.51% accuracy, 75.75% sensitivity, 75.25% specificity, 75.75% precision and Matthews Correlation Coefficient (MCC) of 0.51 in blind testing. IRESPred was found to perform better than the only available viral IRES prediction server, VIPS. The IRESPred server is freely available at http://bioinfo.net.in/IRESPred/.

This review presents a holistic view on the translational potential of the interplay between stromal cells and cancer cells. This interplay is currently being employed for the development of promising preclinical and clinical biomarkers, and the design of small molecule inhibitors, antibodies and small RNAs for (combinatorial) cancer treatment options. In addition, nano-carriers, tissue scaffolds and 3-D based matrices are being developed to precisely and safely deliver these compounds.