Protein synthesis in eukaryotes: The growing biological relevance of cap-independent translation initiation

Ribosome recruitment to eukaryotic mRNAs is generally thought to occur by a scanning mechanism, whereby the 40S ribosomal subunit binds in the vicinity of the 5'cap structure of the mRNA and scans until an AUG codon is encountered in an appropriate sequence context. Study of the picornaviruses allowed the characterization of an alternative mechanism of translation initiation. Picornaviruses can initiate translation via an internal ribosome entry segment (IRES), an RNA structure that directly recruits the 40S ribosomal subunits in a cap and 5' end independent fashion. Since its discovery, the notion of IRESs has extended to a number of different virus families and cellular RNAs. This review summarizes features of both cap-dependent and IRES-dependent mechanisms of translation initiation and discusses the role of cis-acting elements, which include the 5'cap, the 5'-untranslated region (UTR) and the poly(A) tail as well as the possible roles of IRESs as part of a cellular stress response mechanism and in the virus replication cycle.

Nucleolar organizer regions in a model of cell hyperactivity and regression

Nucleolar organizer regions stained with colloidal silver techniques (AgNOR) evidence sites of active rRNA transcription. It has been proved that AgNOR undergo a rise in number and variations in size and shape in conditions which traditionally involve enhanced cell proliferation and rRNA transcription. AgNOR have been described as a marker of malignant transformation in multiple entities. Our laboratory has previously described their value as markers of radioinduced damage. The finding, at light microscopy level, that silver staining persisted at later post-irradiation times when cells are characteristically inactive, prompted the present study to correlate findings at light microscopy level with the ultrastructural analysis of nucleoli and their AgNOR in a model of irradiated skin. We herein attempt to explain the biological significance of AgNOR variations in the different phases of radioinduced response (which involves cellular hyperactivity followed by regressive features). Ten Wistar rats were submitted to local irradiation of the left leg (the shielded right leg was used as control) with 50 Gy x rays and killed 15 days post- irradiation. Silver staining was performed on ultrathin sections. In the basal layer of control epithelium silver affinity was established for fibrillar centers (FC) and fibrillar dense components (DFC). During the phase of radioinduced hyperplasia (1-3 days post-exposure) basal cells exhibit large reticular nucleoli, with irregular contours and silver staining on DFC. In the regressive phase (4-5 days post-irradiation) silver staining persists despite the halt in transcriptional activity, associated to homogeneous and compact nucleoli. These findings suggest caution in the interpretation of silver staining patterns.

Reduction of the accumulation of viral mRNA and of cellular rRNA synthesis in MHV3-infected macrophages of resistant mice

1. After MHV3 infection, only macrophages from resistant A/J mice partially restricted virus growth compared to those from susceptible BALB/c mice (2 logs of difference in virus titer). 2. Cellular ribosomal ribonucleic acid (rRNA) synthesis by MHV3-infected macrophages was decreased only in A/J mouse macrophages as indicated by accumulation of the 28S rRNA fraction. 3. The accumulation of viral messenger ribonucleic acids (mRNAs) in MHV3-infected macrophages was also reduced in A/J mouse macrophages compared to BALB/c mice. 4. In pulse-chase experiments of viral protein synthesis, the appearance, glycosylation and cleavage of glycoprotein S, as well as the metabolism of nucleoprotein N were delayed in A/J mouse macrophages. 5. These data show that MHV3 infection of A/J mouse macrophages induced an imbalanced accumulation of the 28S fraction of rRNA. Furthermore the synthesis of mRNAs correlated with viral protein synthesis in both A/J and BALB/c macrophages, but was delayed in A/J mice. 6. These results suggest that the partial restriction of MHV3 replication in macrophages of resistant A/J mice may take place during or before the mRNA synthesis, although it is correlated with the appearance, glycosylation, cleavage and metabolism of viral proteins