The global translation profile in a ribosomal protein mutant resembles that of an eIF3 mutant

Background

Genome-wide assays performed in Arabidopsis and other organisms have revealed that the translation status of mRNAs responds dramatically to different environmental stresses and genetic lesions in the translation apparatus. To identify additional features of the global landscape of translational control, we used microarray analysis of polysomal as well as non-polysomal mRNAs to examine the defects in translation in a poly(A) binding protein mutant, pab2 pab8, as well as in a mutant of a large ribosomal subunit protein, rpl24b/shortvalve1.

Results

The mutation of RPL24B stimulated the ribosome occupancy of mRNAs for nuclear encoded ribosomal proteins. Detailed analysis yielded new insights into the translational regulon containing the ribosomal protein mRNAs. First, the ribosome occupancy defects in the rpl24b mutant partially overlapped with those in a previously analyzed initiation factor mutant, eif3h. Second, a group of mRNAs with incomplete coding sequences appeared to be uncoupled from the regulon, since their dependence on RPL24B differed from regular mRNAs. Third, different sister paralogs of the ribosomal proteins differed in their translation state in the wild-type. Some sister paralogs also differed in their response to the rpl24b mutation. In contrast to rpl24b, the pab2 pab8 mutant revealed few gene specific translational defects, but a group of seed storage protein mRNAs were stimulated in their ribosome occupancy. In the course of this work, while optimizing the statistical analysis of ribosome occupancy data, we collected 12 biological replicates of translation states from wild-type seedlings. We defined 20% of mRNAs as having a high variance in their translation state. Many of these mRNAs were functionally associated with responses to the environment, suggesting that subtle variation in the environmental conditions is sensed by plants and transduced to affect the translational efficiency of hundreds of mRNAs.

Conclusions

These data represent the first genome-wide analysis of translation in a eukaryote defective in the large ribosomal subunit. RPL24 and eIF3h play similar but non-identical roles in eukaryotic translation. The data also shed light on the fine structure of the regulon of ribosomal protein mRNAs.

     

lemmingA encodes the Apc11 subunit of the APC/C in Drosophila melanogaster that forms a ternary complex with the E2-C type ubiquitin conjugating enzyme, Vihar and Morula/Apc2

Background

The spindle assembly checkpoint (SAC) inhibits anaphase progression in the presence of insufficient kinetochore-microtubule attachments, but cells can eventually override mitotic arrest by a process known as mitotic slippage or adaptation. This is a problem for cancer chemotherapy using microtubule poisons.

Results

Here we describe mitotic slippage in yeast bub2?? mutant cells that are defective in the repression of precocious telophase onset (mitotic exit). Precocious activation of anaphase promoting complex/cyclosome (APC/C)-Cdh1 caused mitotic slippage in the presence of nocodazole, while the SAC was still active. APC/C-Cdh1, but not APC/C-Cdc20, triggered anaphase progression (securin degradation, separase-mediated cohesin cleavage, sister-chromatid separation and chromosome missegregation), in addition to telophase onset (mitotic exit), during mitotic slippage. This demonstrates that an inhibitory system not only of APC/C-Cdc20 but also of APC/C-Cdh1 is critical for accurate chromosome segregation in the presence of insufficient kinetochore-microtubule attachments.

Conclusions

The sequential activation of APC/C-Cdc20 to APC/C-Cdh1 during mitosis is central to accurate mitosis. Precocious activation of APC/C-Cdh1 in metaphase (pre-anaphase) causes mitotic slippage in SAC-activated cells. For the prevention of mitotic slippage, concomitant inhibition of APC/C-Cdh1 may be effective for tumor therapy with mitotic spindle poisons in humans.     

High-Sensitivity and High-Resolution In Situ Hybridization of Coding and Long Non-coding RNAs in Vertebrate Ovaries and Testes

Background

Subcellular localization of coding and non-coding RNAs has emerged as major regulatory mechanisms of gene expression in various cell types and many organisms. However, techniques that enable detection of the subcellular distribution of these RNAs with high sensitivity and high resolution remain limited, particularly in vertebrate adult tissues and organs. In this study, we examined the expression and localization of mRNAs encoding Pou5f1/Oct4, Mos, Cyclin B1 and Deleted in Azoospermia-like (Dazl) in zebrafish and mouse ovaries by combining tyramide signal amplification (TSA)-based in situ hybridization with paraffin sections which can preserve cell morphology of tissues and organs at subcellular levels. In addition, the distribution of a long non-coding RNA (lncRNA), lncRNA-HSVIII, in mouse testes was examined by the same method.

Results

The mRNAs encoding Mos, Cyclin B1 and Dazl were found to assemble into distinct granules that were distributed in different subcellular regions of zebrafish and mouse oocytes, suggesting conserved and specific regulations of these mRNAs. The lncRNA-HSVIII was first detected in the nucleus of spermatocytes at prophase I of the meiotic cell cycle and was then found in the cytoplasm of round spermatids, revealing expression patterns of lncRNA during germ cell development. Collectively, the in situ hybridization method demonstrated in this study achieved the detection and comparison of precise distribution patterns of coding and non-coding RNAs at subcellular levels in single cells of adult tissues and organs.

Conclusions

This high-sensitivity and high-resolution in situ hybridization is applicable to many vertebrate species and to various tissues and organs and will be useful for studies on the subcellular regulation of gene expression at the level of RNA localization.

     

Genetic variations regulate alternative splicing in the 5' untranslated regions of the mouse glioma-associated oncogene 1, Gli1

Background  

Alternative splicing is one of the key mechanisms that generate biological diversity. Even though alternative splicing also occurs in the 5' and 3' untranslated regions (UTRs) of mRNAs, the understanding of the significance and the regulation of these variations is rather limited.     

Vision is adapted to the natural level of blur present in the retinal image

Background

The image formed by the eye''s optics is inherently blurred by aberrations specific to an individual''s eyes. We examined how visual coding is adapted to the optical quality of the eye.

Methods and Findings

We assessed the relationship between perceived blur and the retinal image blur resulting from high order aberrations in an individual''s optics. Observers judged perceptual blur in a psychophysical two-alternative forced choice paradigm, on stimuli viewed through perfectly corrected optics (using a deformable mirror to compensate for the individual''s aberrations). Realistic blur of different amounts and forms was computer simulated using real aberrations from a population. The blur levels perceived as best focused were close to the levels predicted by an individual''s high order aberrations over a wide range of blur magnitudes, and were systematically biased when observers were instead adapted to the blur reproduced from a different observer''s eye.

Conclusions

Our results provide strong evidence that spatial vision is calibrated for the specific blur levels present in each individual''s retinal image and that this adaptation at least partly reflects how spatial sensitivity is normalized in the neural coding of blur.     

Reprogramming the genetic code: The emerging role of ribosomal frameshifting in regulating cellular gene expression

Reading frame maintenance is a critical property of ribosomes. However, a number of genetic elements have been described that can induce ribosomes to shift on mRNAs, the most well understood of which are a class that directs ribosomal slippage by one base in 5' (‐1) direction. This is referred to as programmed ‐1 ribosomal frameshifting (‐1 PRF). Recently, a new ‐1 PRF promoting element was serendipitously discovered in a study examining the effects of stretches of adenosines in the coding sequences of mRNAs. Here, we discuss this finding, recent studies describing how ‐1 PRF is used to control gene expression in eukaryotes, and how ‐1 PRF is itself regulated. The implications of dysregulation of ‐1 PRF on human health are examined, as are possible new areas in which novel ‐1 PRF promoting elements might be discovered. Also watch the Video Abstract .

     

Genome-Wide Analyses of Radioresistance-Associated miRNA Expression Profile in Nasopharyngeal Carcinoma Using Next Generation Deep Sequencing

Background

Rapidly growing evidence suggests that microRNAs (miRNAs) are involved in a wide range of cancer malignant behaviours including radioresistance. Therefore, the present study was designed to investigate miRNA expression patterns associated with radioresistance in NPC.

Methods

The differential expression profiles of miRNAs and mRNAs associated with NPC radioresistance were constructed. The predicted target mRNAs of miRNAs and their enriched signaling pathways were analyzed via biological informatical algorithms. Finally, partial miRNAs and pathways-correlated target mRNAs were validated in two NPC radioreisitant cell models.

Results

50 known and 9 novel miRNAs with significant difference were identified, and their target mRNAs were narrowed down to 53 nasopharyngeal-/NPC-specific mRNAs. Subsequent KEGG analyses demonstrated that the 53 mRNAs were enriched in 37 signaling pathways. Further qRT-PCR assays confirmed 3 down-regulated miRNAs (miR-324-3p, miR-93-3p and miR-4501), 3 up-regulated miRNAs (miR-371a-5p, miR-34c-5p and miR-1323) and 2 novel miRNAs. Additionally, corresponding alterations of pathways-correlated target mRNAs were observed including 5 up-regulated mRNAs (ICAM1, WNT2B, MYC, HLA-F and TGF-β1) and 3 down-regulated mRNAs (CDH1, PTENP1 and HSP90AA1).

Conclusions

Our study provides an overview of miRNA expression profile and the interactions between miRNA and their target mRNAs, which will deepen our understanding of the important roles of miRNAs in NPC radioresistance.