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Maximizing light capture by light-harvesting pigment optimization represents an attractive but challenging strategy to improve photosynthetic efficiency. Here, we report that loss of a previously uncharacterized gene, HIGH PHOTOSYNTHETIC EFFICIENCY1 (HPE1), optimizes light-harvesting pigments, leading to improved photosynthetic efficiency and biomass production. Arabidopsis (Arabidopsis thaliana) hpe1 mutants show faster electron transport and increased contents of carbohydrates. HPE1 encodes a chloroplast protein containing an RNA recognition motif that directly associates with and regulates the splicing of target RNAs of plastid genes. HPE1 also interacts with other plastid RNA-splicing factors, including CAF1 and OTP51, which share common targets with HPE1. Deficiency of HPE1 alters the expression of nucleus-encoded chlorophyll-related genes, probably through plastid-to-nucleus signaling, causing decreased total content of chlorophyll (a+b) in a limited range but increased chlorophyll a/b ratio. Interestingly, this adjustment of light-harvesting pigment reduces antenna size, improves light capture, decreases energy loss, mitigates photodamage, and enhances photosynthetic quantum yield during photosynthesis. Our findings suggest a novel strategy to optimize light-harvesting pigments that improves photosynthetic efficiency and biomass production in higher plants.

The linker of nucleoskeleton and cytoskeleton (LINC) complex is a multifunctional protein complex that is involved in various processes at the nuclear envelope, including nuclear migration, mechanotransduction, chromatin tethering and DNA damage response. We recently showed that a nuclear envelope protein, Sad1 and UNC84 domain protein 1 (SUN1), a component of the LINC complex, has a critical function in cell migration. Although ionizing radiation activates cell migration and invasion in vivo and in vitro, the underlying molecular mechanism remains unknown. Here, we examined the involvement of the LINC complex in radiation‐enhanced cell migration and invasion. A sublethal dose of X‐ray radiation promoted human breast cancer MDA‐MB‐231 cell migration and invasion, whereas carbon ion beam radiation suppressed these processes in a dose‐dependent manner. Depletion of SUN1 and SUN2 significantly suppressed X‐ray‐enhanced cell migration and invasion. Moreover, depletion or overexpression of each SUN1 splicing variant revealed that SUN1_888 containing 888 amino acids of SUN1 but not SUN1_916 was required for X‐ray‐enhanced migration and invasion. In addition, the results suggested that X‐ray irradiation affected the expression level of SUN1 splicing variants and a SUN protein binding partner, nesprins. Taken together, our findings supported that the LINC complex contributed to photon‐enhanced cell migration and invasion. The LINC complex is a multifunctional protein complex. This study showed that the LINC complex contributed to photon‐enhanced cancer cell migration.

RNA splicing factors are essential for the viability of all eukaryotic cells; however, in metazoans some cell types are exquisitely sensitive to disruption of splicing factors. Neuronal cells represent one such cell type, and defects in RNA splicing factors can lead to neurodegenerative diseases. The basis for this tissue selectivity is not well understood owing to difficulties in analyzing the consequences of splicing factor defects in whole-animal systems. Here, we use zebrafish mutants to show that loss of spliceosomal components, including splicing factor 3b, subunit 1 (sf3b1), causes increased DNA double-strand breaks and apoptosis in embryonic neurons. Moreover, these mutants show a concomitant accumulation of R-loops, which are non-canonical nucleic acid structures that promote genomic instability. Dampening R-loop formation by conditional induction of ribonuclease H1 in sf3b1 mutants reduced neuronal DNA damage and apoptosis. These findings show that splicing factor dysfunction leads to R-loop accumulation and DNA damage that sensitizes embryonic neurons to apoptosis. Our results suggest that diseases associated with splicing factor mutations could be susceptible to treatments that modulate R-loop levels.

Accurate assessment of the individual exposure dose based on easily accessible samples (e.g. blood) immediately following a radiological accident is crucial. We aimed at developing a robust transcription-based signature for biodosimetry from human peripheral blood mononuclear cells irradiated with different doses of X-rays (0.1 and 1.0 Gy) at a dose rate of 0.26 Gy/min. Genome-wide radiation-induced changes in mRNA expression were evaluated at both gene and exon level. Using exon-specific qRT-PCR, we confirmed that several biomarker genes are alternatively spliced or transcribed after irradiation and that different exons of these genes exhibit significantly different levels of induction. Moreover, a significant number of radiation-responsive genes were found to be genomic neighbors. Using three different classification models we found that gene and exon signatures performed equally well on dose prediction, as long as more than 10 features are included. Together, our results highlight the necessity of evaluating gene expression at the level of single exons for radiation biodosimetry in particular and transcriptional biomarker research in general. This approach is especially advisable for practical gene expression-based biodosimetry, for which primer- or probe-based techniques would be the method of choice.

The P2X7 receptor (P2X7-R) is expressed in the retina and brain and has been implicated in neurodegenerative diseases. However, whether it is expressed by neurons and plays a role as a neurotransmitter receptor has been the subject of controversy. In this study, we first show that the novel vesicular transporter for ATP, VNUT, is expressed in the retina, verifying the presence of the molecular machinery for ATP to act as neurotransmitter at P2X7-Rs. Secondly we show the presence of P2X7-R mRNA and protein in the retina and cortex and absence of the full length variant 1 of the receptor in the P2X7-R knock out (P2X7-KO) mouse. The role of the P2X7-R in neuronal function of the retina was assessed by comparing the electroretinogram response of P2X7-KO with WT mice. The rod photoreceptor response was found to be similar, while both rod and cone pathway post-photoreceptor responses were significantly larger in P2X7-KO mice. This suggests that activation of P2X7-Rs modulates output of second order retinal neurons. In line with this finding, P2X7-Rs were found in the outer plexiform layer and on inner retinal cell classes, including horizontal, amacrine and ganglion cells. The receptor co-localized with conventional synapses in the IPL and was expressed on amacrine cells post-synaptic to rod bipolar ribbon synapses. In view of the changes in visual function in the P2X7-KO mouse and the immunocytochemical location of the receptor in the normal retina, it is likely the P2X7-R provides excitatory input to photoreceptor terminals or to inhibitory cells that shape both the rod and cone pathway response.

Despite a strict conservation of the vascular tissues in vascular plants (tracheophytes), our understanding of the genetic basis underlying the differentiation of secondary cell wall-containing cells in the xylem of tracheophytes is still far from complete. Using coexpression analysis and phylogenetic conservation across sequenced tracheophyte genomes, we identified a number of Arabidopsis (Arabidopsis thaliana) genes of unknown function whose expression is correlated with secondary cell wall deposition. Among these, the Arabidopsis VASCULAR-RELATED UNKNOWN PROTEIN1 (VUP1) gene encodes a predicted protein of 24 kD with no annotated functional domains but containing domains that are highly conserved in tracheophytes. Here, we show that the VUP1 expression pattern, determined by promoter-ß-glucuronidase reporter gene expression, is associated with vascular tissues, while vupl loss-of-function mutants exhibit collapsed morphology of xylem vessel cells. Constitutive overexpression of VUP1 caused dramatic and pleiotropic developmental defects, including severe dwarfism, dark green leaves, reduced apical dominance, and altered photomorphogenesis, resembling brassinosteroid-deficient mutants. Constitutive overexpression of VUP homologs from multiple tracheophyte species induced similar defects. Whole-genome transoiptome analysis revealed that overexpression of VUP1 represses the expression of many brassinosteroid-and auxin-responsive genes. Additionally, deletion constructs and site-directed mutagenesis were used to identify critical domains and amino acids required for VUP1 function. Altogether, our data suggest a conserved role for VUP1 in regulating secondary wall formation during vascular development by tissue-or cell-specific modulation of hormone signaling pathways.

UV radiation is the most important environmental skin aggressor, causing cancer and other problems. This paper reports the use of oligonucleotide microarray technology to determine changes in gene expression in human keratinocytes after UVB treatment. Examination of the effects of different doses at different times after irradiation gave a global picture of the keratinocyte response to this type of insult. Five hundred thirty-nine regulated transcripts were found and organized into nine different clusters depending on behavior patterns. Classification of these genes into 23 functional categories revealed that several biological processes are globally affected by UVB. In addition to confirming a majority up-regulation of the transcripts related to the UV-specific inflammatory and stress responses, significant increases were seen in the expression of genes involved in basal transcription, splicing, and translation as well as in the proteasome-mediated degradation category. On the other hand, those transcripts belonging to the metabolism and adhesion categories were strongly down-regulated. These results demonstrate the complexity of the transcriptional profile of the UVB response, describe several cellular processes previously not known to be affected by UV irradiation, and serve as a basis for the global characterization of UV-regulated genes and pathways.

A rare class of introns with AT-AC at their termini recently has been identified in metazoan genes. Splicing of these introns requires a different set of small nuclear ribonucleoprotein particles (snRNPs) (U11, U12, U5, and U4atac/U6atac) compared with the snRNPs (U1, U2, U5, and U4/U6) required for splicing the majority of pre-mRNA introns, but otherwise little is known regarding the excision of AT-AC introns. Here we use site-specific 4-thiouridine (4SU) crosslinking analysis to dissect the mechanism of 5′ splice site recognition during in vitro splicing of the AT-AC intron from the P120 pre-mRNA. Upon irradiation with 365-nm UV light, three P120 substrates, each with a single 4SU substitution near the 5′ splice site (at position +2, +4, or +7), produce two early ATP-independent crosslinks with similar kinetics. For one of the substrates, P120-4SU+2, a third ATP-requiring crosslink forms as the two early crosslinks diminish. RNase H digestion coupled with Northern blotting indicates that the two early crosslinks generated with P120-4SU+2 contain the U11 small nuclear RNA. Reverse transcription-PCR followed by cloning and sequencing demonstrates that the third crosslink involves U6atac. The dynamic appearance of the three crosslinks correlates with the kinetics of the splicing reaction and suggests that the 5′ splice site is recognized first by U11 and then by U6atac. Our results argue that the splicing of AT-AC introns is mechanistically similar to the splicing of the major class of introns and that the U11 and U6atac snRNPs in the AT-AC spliceosome fulfill analogous roles to U1 and U6, respectively, in the major spliceosome.

A mammalian protein of approximately 220 kDa (p220) was UV-crosslinked to precursor mRNAs (pre-mRNAs) under splicing conditions. The kinetics and biochemical requirements of the UV-crosslinking of p220 corresponded to the kinetics and biochemical requirements of spliceosome formation. On Western blots, antibodies against the yeast splicing factor PRP8 recognized a doublet of proteins, the faster migrating of which comigrated with p220. Furthermore, UV-crosslinked p220 was immunoprecipitated with anti-PRP8 antisera. These results suggest structural conservation of the splicing factor PRP8 from yeast to mammals and show that this protein is in close proximity to the pre-mRNA in the spliceosome

Cervical cancer (CCa) is the fourth leading cause of cancer-related deaths among women worldwide, with nearly 90% of cases in low- and middle-income countries, especially in Sub-Saharan Africa. This study explores the roles of circular ribonucleic acids (circRNAs), hsa_circ_0001038 and circRNA_400029, and the impact of the serine/arginine-rich splicing factor 3 (SRSF3) inhibitor, theophylline, in CCa cell lines. We utilized cell cycle fluorescence-activated cell sorting (FACS) and Annexin V/propidium iodide (PI) assays to evaluate theophylline’s effects on SiHa and C33A cell lines. Results showed S-phase arrest in SiHa and G2/M arrest in C33A, with significant cytotoxic effects indicated by apoptosis analysis. Using CircAtlas, we identified micro ribonucleic acids (miRNAs) binding to hsa_circ_0001038, particularly miR-205-5p, which has a tumour-suppressive role. miRTarBase identified miR-16-5p as a key interacting miRNA for circRNA_400029. We constructed a competing endogenous ribonucleic acid (ceRNA) network, revealing multiple miRNA targets. Pathway analysis via the Kyoto Encyclopedia of Genes and Genomes (KEGG) highlighted critical signalling pathways involved in CCa oncogenesis. In conclusion, theophylline demonstrates cytotoxicity in CCa cells, suggesting its potential for repurposing in CCa theranostics, though further optimization is necessary.