Emerging links between hypermutation of antibody genes and DNA polymerases

Substantial antibody variability is created when nucleotide substitutions are introduced into immunoglobulin variable genes by a controlled process of hypermutation. Evidence points to a mechanism involving DNA repair events at sites of targeted breaks. In vertebrate cells, there are many recently identified DNA polymerases that inaccurately copy templates. Some of these are candidates for enzymes that introduce base changes during hypermutation. Recent research has focused on possible roles for DNA polymerases zeta (POLZ), eta (POLH), iota (POLI), and mu (POLM) in the process.     

Emerging links between the biological clock and the DNA damage response

For life forms to survive, they must adapt to their environmental conditions. One such factor that impacts on both prokaryotic and eukaryotic organisms is the light–dark cycle, a consequence of planetary rotation in relation to our sun. In mammals, the daily light cycle has affected the regulation of many cellular processes such as sleep–wake and calorific intake activities, hormone secretion, blood pressure and immune system responses. Such rhythmic behaviour is the consequence of circadian rhythm/biological clock (BC) systems which are controlled in a light stimulus-dependent manner by a master clock called the suprachiasmatic nucleus (SCN) situated within the anterior hypothalamus. Peripheral clocks located in other organs such as the liver and kidneys relay signals from the SCN, which ultimately leads to tightly controlled expression of several protein families that in turn act on a broad range of cellular functions. Work in lower organisms has demonstrated a link between aging processes and BC factors, and studies in both animal models and clinical trials have postulated a role for certain BC-associated proteins in tumourigenesis and cancer progression. Recent exciting data reported within the last year or so have now established a molecular link between specific BC proteins and factors that control the mammalian cell cycle and DNA damage checkpoints. This mini review will focus on these discoveries and emphasise how such BC proteins may be involved, through their interplay with cell cycle/DNA damage response pathways, in the development of human disease such as cancer.     

Conflict between translation initiation and elongation in vertebrate mitochondrial genomes

The strand-biased mutation spectrum in vertebrate mitochondrial genomes results in an AC-rich L-strand and a GT-rich H-strand. Because the L-strand is the sense strand of 12 protein-coding genes out of the 13, the third codon position is overall strongly AC-biased. The wobble site of the anticodon of the 22 mitochondrial tRNAs is either U or G to pair with the most abundant synonymous codon, with only one exception. The wobble site of Met-tRNA is C instead of U, forming the Watson-Crick match with AUG instead of AUA, the latter being much more frequent than the former. This has been attributed to a compromise between translation initiation and elongation; i.e., AUG is not only a methionine codon, but also an initiation codon, and an anticodon matching AUG will increase the initiation rate. However, such an anticodon would impose selection against the use of AUA codons because AUA needs to be wobble-translated. According to this translation conflict hypothesis, AUA should be used relatively less frequently compared to UUA in the UUR codon family. A comprehensive analysis of mitochondrial genomes from a variety of vertebrate species revealed a general deficiency of AUA codons relative to UUA codons. In contrast, urochordate mitochondrial genomes with two tRNA(Met) genes with CAU and UAU anticodons exhibit increased AUA codon usage. Furthermore, six bivalve mitochondrial genomes with both of their tRNA-Met genes with a CAU anticodon have reduced AUA usage relative to three other bivalve mitochondrial genomes with one of their two tRNA-Met genes having a CAU anticodon and the other having a UAU anticodon. We conclude that the translation conflict hypothesis is empirically supported, and our results highlight the fine details of selection in shaping molecular evolution.     

Bypassing translation initiation

A high-resolution cryo-EM reconstruction of a ribosome-bound dicistrovirus IRES (Schüler et al., 2006) and the crystal structure of its ribosome binding domain (Pfingsten et al., 2006) provide new insights into an exceptional eukaryotic translation mechanism.     

Sulfolobus solfataricus translation initiation factor 1 stimulates translation initiation complex formation

The eukaryotic translation initiation factor 1 binds to the ribosome during translation initiation. It is instrumental for initiator-tRNA and mRNA binding, and has a function in selection of the authentic start codon. Here, we show that the archaeal homolog aIF1 has analogous functions. The aIF1 protein of the archaeon Sulfolobus solfataricus is bound to the small ribosomal subunit during translation initiation and accelerates binding of initiator-tRNA and mRNA to the ribosome. Accordingly, aIF1 stimulated translation of an mRNA in a S. solfataricus in vitro translation system. Moreover, this study suggested that the C terminus of the factor is of relevance for its function.     

Emerging roles of chloride channels in human diseases

In the past decade, there has been remarkable progress in understanding of the roles of Cl(-) channels in the development of human diseases. Genetic studies in humans have identified mutations in the genes encoding Cl(-) channels which lead to a loss of Cl(-) channel activity. These mutations are responsible for the development of a variety of deleterious diseases in muscle, kidney, bone and brain including myotonia congenita, dystrophia myotonica, cystic fibrosis, osteopetrosis and epilepsy. Recent studies indicate that some diseases may develop as a result of Cl(-) channel activation. There is growing evidence that the progression of glioma in the brain and the growth of the malaria parasite in red blood cells may be mediated through Cl(-) channel activation. These findings suggest that Cl(-) channels may be novel targets for the pharmacological treatment of a broad spectrum of diseases. This review discusses the proposed roles of abnormal Cl(-) channel activity in the pathogenesis of human diseases.     

Glucocorticoids modulate amino acid-induced translation initiation in human skeletal muscle

Amino acids are unique anabolic agents in that they nutritively signal to mRNA translation initiation and serve as substrates for protein synthesis in skeletal muscle. Glucocorticoid excess antagonizes the anabolic action of amino acids on protein synthesis in laboratory animals. To examine whether excessive glucocorticoids modulate mixed amino acid-signaled translation initiation in human skeletal muscle, we infused an amino acid mixture (10% Travasol) systemically to 16 young healthy male volunteers for 6 h in the absence (n = 8) or presence (n = 8) of glucocorticoid excess (dexamethasone 2 mg orally every 6 h for 3 days). Vastus lateralis muscles were biopsied before and after amino acid infusion, and the phosphorylation of eukaryotic initiation factor (eIF) 4E-binding protein 1 (4E-BP1), ribosomal protein S6 kinase (p70(S6K)), and eIF2alpha and the guanine nucleotide exchange activity of eIF2B were measured. Systemic infusion of mixed amino acids significantly stimulated the phosphorylation of 4E-BP1 (P < 0.04) and p70(S6K) (P < 0.001) and the dephosphorylation of eIF2alpha (P < 0.003) in the control group. Dexamethasone treatment did not alter the basal phosphorylation state of 4E-BP1, p70(S6K), or eIF2alpha; however, it abrogated the stimulatory effect of amino acid infusion on the phosphorylation of 4E-BP1 (P = 0.31) without affecting amino acid-induced phosphorylation of p70(S6K) (P = 0.002) or dephosphorylation of eIF2alpha (P = 0.003). Neither amino acid nor dexamethasone treatment altered the guanine nucleotide exchange activity of eIF2B. We conclude that changes of amino acid concentrations within the physiological range stimulate mRNA translation by enhancing the binding of mRNA to the 43S preinitiation complex, and the activity of p70(S6K) and glucocorticoid excess blocks the former action in vivo in human skeletal muscle.     

Ubiquitination and proteasome-dependent degradation of human eukaryotic translation initiation factor 4E

Translation initiation factor 4E (eIF4E) is a cytoplasmic cap-binding protein that is required for cap-dependent translation initiation. Here, we have shown that eIF4E is ubiquitinated primarily at Lys-159 and incubation of cells with a proteasome inhibitor leads to increased eIF4E levels, suggesting the proteasome-dependent proteolysis of ubiquitinated eIF4E. Ubiquitinated eIF4E retained its cap binding ability, whereas eIF4E phosphorylation and eIF4G binding were reduced by ubiquitination. The W73A mutant of eIF4E exhibited enhanced ubiquitination/degradation, and 4E-BP overexpression protected eIF4E from ubiquitination/degradation. Because heat shock or the expression of the carboxyl terminus of heat shock cognate protein 70-interacting protein (Chip) dramatically increased eIF4E ubiquitination, Chip may be at least one ubiquitin E3 ligase responsible for eIF4E ubiquitination.     

Eukaryotic translation initiation factors and regulators

Significant progress has been made over the past several years on structural studies of the eukaryotic translation initiation factors that facilitate the assembly of a translation-competent ribosome at the initiation codon of an mRNA. These structural studies have revealed the repeated use of a set of common structural folds, highlighted the evolutionary conservation of the translation apparatus, and provided insight into the mechanism and regulation of cellular and viral protein synthesis.     

Calicivirus translation initiation requires an interaction between VPg and eIF 4 E

Unlike other positive-stranded RNA viruses that use either a 5'-cap structure or an internal ribosome entry site to direct translation of their messenger RNA, calicivirus translation is dependent on the presence of a protein covalently linked to the 5' end of the viral genome (VPg). We have shown a direct interaction of the calicivirus VPg with the cap-binding protein eIF 4 E. This interaction is required for calicivirus mRNA translation, as sequestration of eIF 4 E by 4 E-BP 1 inhibits translation. Functional analysis has shown that VPg does not interfere with the interaction between eIF 4 E and the cap structure or 4 E-BP 1, suggesting that VPg binds to eIF 4 E at a different site from both cap and 4 E-BP 1. This work lends support to the idea that calicivirus VPg acts as a novel 'cap substitute' during initiation of translation on virus mRNA.     

Emerging therapeutics targeting mRNA translation

A defining feature of many cancers is deregulated translational control. Typically, this occurs at the level of recruitment of the 40S ribosomes to the 5'-cap of cellular messenger RNAs (mRNAs), the rate-limiting step of protein synthesis, which is controlled by the heterotrimeric eukaryotic initiation complex eIF4F. Thus, eIF4F in particular, and translation initiation in general, represent an exploitable vulnerability and unique opportunity for therapeutic intervention in many transformed cells. In this article, we discuss the development, mode of action and biological activity of a number of small-molecule inhibitors that interrupt PI3K/mTOR signaling control of eIF4F assembly, as well as compounds that more directly block eIF4F activity.     

Genomic context analysis in Archaea suggests previously unrecognized links between DNA replication and translation

Background  

Comparative analysis of genomes is valuable to explore evolution of genomes, deduce gene functions, or predict functional linking between proteins. Here, we have systematically analyzed the genomic environment of all known DNA replication genes in 27 archaeal genomes to infer new connections for DNA replication proteins from conserved genomic associations.     

Coupled folding during translation initiation

The structure of the eukaryotic initiation factor eIF4E bound to a cognate domain of eIF4G and m(7)GDP in this issue of Cell shows that these factors undergo coupled folding to form a stable complex with high cap binding activity that promotes efficient ribosomal attachment to mRNA during translation initiation.     

Eukaryotic translation initiation factor 4E availability controls the switch between cap-dependent and internal ribosomal entry site-mediated translation

Translation of m7G-capped cellular mRNAs is initiated by recruitment of ribosomes to the 5' end of mRNAs via eukaryotic translation initiation factor 4F (eIF4F), a heterotrimeric complex comprised of a cap-binding subunit (eIF4E) and an RNA helicase (eIF4A) bridged by a scaffolding molecule (eIF4G). Internal translation initiation bypasses the requirement for the cap and eIF4E and occurs on viral and cellular mRNAs containing internal ribosomal entry sites (IRESs). Here we demonstrate that eIF4E availability plays a critical role in the switch from cap-dependent to IRES-mediated translation in picornavirus-infected cells. When both capped and IRES-containing mRNAs are present (as in intact cells or in vitro translation extracts), a decrease in the amount of eIF4E associated with the eIF4F complex elicits a striking increase in IRES-mediated viral mRNA translation. This effect is not observed in translation extracts depleted of capped mRNAs, indicating that capped mRNAs compete with IRES-containing mRNAs for translation. These data explain numerous reported observations where viral mRNAs are preferentially translated during infection.     

A functional link between Disrupted-In-Schizophrenia 1 and the eukaryotic translation initiation factor 3

Disrupted-In-Schizophrenia 1 (DISC1) was identified as a candidate gene for schizophrenia. DISC1 is disrupted by a balanced t(1;11)(q42.1;q14.3) translocation segregating with schizophrenia and related psychiatric illness in a large Scottish family. Here, we show that DISC1 interacts via its globular domain with the p40 subunit of the eukaryotic translation initiation factor 3. Furthermore, we found that overexpression of DISC1 in SH-SY5Y cells induces the assembly of eIF3- and TIA-1-positive stress granules (SGs), discrete cytoplasmic granules formed in response to environmental stresses. Our findings suggest that DISC1 may function as a translational regulator and may be involved in stress response.