Chloroplasts are unique organelles that not only provide sites for photosynthesis and many metabolic processes, but also are sensitive to various environmental stresses. Chloroplast proteins are encoded by genes from both nuclear and chloroplast genomes. During chloroplast development and responses to stresses, the robust protein quality control systems are essential for regulation of protein homeostasis and the integrity of chloroplast proteome. In this review, we summarize the regulatory mechanisms of chloroplast protein degradation refer to protease system, ubiquitin-proteasome system, and the chloroplast autophagy. These mechanisms symbiotically play a vital role in chloroplast development and photosynthesis under both normal or stress conditions.

As sessile organisms, plants have evolved sophisticated mechanisms to optimize their growth and development in response to fluctuating nutrient levels. Brassinosteroids (BRs) are a group of plant steroid hormones that play critical roles in plant growth and developmental processes as well as plant responses to environmental stimuli. Recently, multiple molecular mechanisms have been proposed to explain the integration of BRs with different nutrient signaling processes to coordinate gene expression, metabolism, growth, and survival. Here, we review recent advances in understanding the molecular regulatory mechanisms of the BR signaling pathway and the multifaceted roles of BR in the intertwined sensing, signaling, and metabolic processes of sugar, nitrogen, phosphorus, and iron. Further understanding and exploring these BR-related processes and mechanisms will facilitate advances in crop breeding for higher resource efficiency.

Since the discovery of the first transposon by Dr. Barbara McClintock, the prevalence and diversity of transposable elements (TEs) have been gradually recognized. As fundamental genetic components, TEs drive organismal evolution not only by contributing functional sequences (e.g., regulatory elements or "controllers" as phrased by Dr. McClintock) but also by shuffling genomic sequences. In the latter respect, TE-mediated gene duplications have contributed to the origination of new genes and attracted extensive interest. In response to the development of this field, we herein attempt to provide an overview of TE-mediated duplication by focusing on common rules emerging across duplications generated by different TE types. Specifically, despite the huge divergence of transposition machinery across TEs, we identify three common features of various TE-mediated duplication mechanisms, including end bypass, template switching, and recurrent transposition. These three features lead to one common functional outcome, namely, TE-mediated duplicates tend to be subjected to exon shuffling and neofunctionalization. Therefore, the intrinsic properties of the mutational mechanism constrain the evolutionary trajectories of these duplicates. We finally discuss the future of this field including an in-depth characterization of both the duplication mechanisms and functions of TE-mediated duplicates.

Enhancer promoter interaction (EPI) involves most of gene transcriptional regulation in the high eukaryotes. Predicting the EPIs from given genomic loci or DNA sequences is not a trivial task. The benchmarking work so far for EPI predictors is more or less empirical and lacks quantitative model-based comparisons, posing challenges for molecular biologists to obtain reliable EPI predictions. Here, we present an EPI prediction platform, namely Delta.EPI. Based on a statistic model of the data integration, Delta.EPI is capable of comprehensively assessing the predictions from four state-of-the-art EPI predictors. Equipped with a user-friendly interface and visualization platform, Delta.EPI presents the sorted results with the confidence of EPI relevance, which may guide the molecular biologists who lack the pre-knowledge of the algorithms of EPI prediction. Last, we showcase the utility of Delta.EPI with a case study. Delta.EPI provides a powerful tool to fuel the gene regulation and 3D genome studies by ease-to-access EPI predictions. Delta.EPI can be freely accessed at https://ngdc.cncb.ac.cn/deltaEPI/.

Ferroptosis, a regulated and iron-dependent form of cell death characterized by peroxidation of membrane phospholipids, has tremendous potential for the therapy of human diseases. The causal link between phospholipid homeostasis and ferroptosis is incompletely understood. Here, we reveal that spin-4, a previously identified regulator of the "B12-one-carbon cycle-phosphatidylcholine (PC)" pathway, sustains germline development and fertility by ensuring PC sufficiency in the nematode Caenorhabditis elegans. Mechanistically, SPIN-4 regulates lysosomal activity which is required for B12-associated PC synthesis. PC deficiency-induced sterility can be rescued by reducing the levels of polyunsaturated fatty acids, reactive oxygen species, and redox-active iron, which indicates that the sterility is mediated by germline ferroptosis. These results highlight the critical role of PC homeostasis in ferroptosis susceptibility and offer a new target for pharmacological approaches.

Gene loss is common and influences genome evolution trajectories. Multiple adaptive strategies to compensate for gene loss have been observed, including copy number gain of paralogous genes and mutations in genes of the same pathway. By using the Ubl-specific protease 2 (ULP2) eviction model, we identify compensatory mutations in the homologous gene ULP1 by laboratory evolution and find that these mutations are capable of rescuing defects caused by the loss of ULP2. Furthermore, bioinformatics analysis of genomes of yeast gene knockout library and natural yeast isolate datasets suggests that point mutations of a homologous gene might be an additional mechanism to compensate for gene loss.

O-GlcNAcylation is a post-translational modification that serves as a cellular nutrient sensor and participates in multiple physiological and pathological processes. However, it remains uncertain whether O-GlcNAcylation is involved in the regulation of phagocytosis. Here, we demonstrate a rapid increase in protein O-GlcNAcylation in response to phagocytotic stimuli. Knockout of the O-GlcNAc transferase or pharmacological inhibition of O-GlcNAcylation dramatically blocks phagocytosis, resulting in the disruption of retinal structure and function. Mechanistic studies reveal that the O-GlcNAc transferase interacts with Ezrin, a membrane-cytoskeleton linker protein, to catalyze its O-GlcNAcylation. Our data further show that Ezrin O-GlcNAcylation promotes its localization to the cell cortex, thereby stimulating the membrane-cytoskeleton interaction needed for efficient phagocytosis. These findings identify a previously unrecognized role for protein O-GlcNAcylation in phagocytosis with important implications in both health and diseases.

Mouse dental papilla cells (mDPCs) are cranial neural crest-derived dental mesenchymal cells that give rise to dentin-secreting odontoblasts after the bell stage during odontogenesis. The odontoblastic differentiation of mDPCs is spatiotemporally regulated by transcription factors (TFs). Our previous work reveals that chromatin accessibility was correlated with the occupation of the basic leucine zipper TF family during odontoblastic differentiation. However, the detailed mechanism by which TFs regulate the initiation of odontoblastic differentiation remains elusive. Here, we report that phosphorylation of ATF2 (p-ATF2) is particularly increased during odontoblastic differentiation in vivo and in vitro. ATAC-seq and p-ATF2 CUT&Tag experiments further demonstrate a high correlation between p-ATF2 localization and increased chromatin accessibility of regions near mineralization-related genes. Knockdown of Atf2 inhibits the odontoblastic differentiation of mDPCs, while overexpression of p-ATF2 promotes odontoblastic differentiation. ATAC-seq after overexpression of p-ATF2 reveals that p-ATF2 increases the chromatin accessibility of regions adjacent to genes associated with matrix mineralization. Furthermore, we find that p-ATF2 physically interacts with and promotes H2BK12 acetylation. Taken together, our findings reveal a mechanism that p-ATF2 promotes odontoblastic differentiation at initiation via remodeling chromatin accessibility and emphasize the role of the phosphoswitch model of TFs in cell fate transitions.

Prime editing (PE) is a versatile genome editing tool without the need for double-stranded DNA breaks or donor DNA templates, but is limited by low editing efficiency. We previously fused the M-MLV reverse transcriptase to the Cas9 nickase, generating the PE2 (v1) system, but the editing efficiency of this system is still low. Here we develop different versions of PE2 by adding the 5'-to-3' exonuclease at different positions of the nCas9-M-MLV RT fusion protein. PE2 (v2), in which the T5 exonuclease fused to the N-terminus of the nCas9-MMLV fusion protein enhances prime editing efficiency of base substitutions, deletions, and insertions at several genomic sites by 1.7- to 2.9-fold in plant cells compared to PE2 (v1). The improved editing efficiency of PE2 (v2) is further confirmed by generating increased heritable prime edits in stable transgenic plants compared to the previously established PE-P1, PE-P2, and PPE systems. Using PE2 (v2), we generate herbicide-resistant rice by simultaneously introducing mutations causing amino acid substitutions at two target sites. The PE efficiency is further improved by combining PE2 (v2) and dual-pegRNAs. Taken together, the increased genome editing efficiency of PE2 (v2) developed in this study may enhance the applications of PE in plants.

The phytohormone auxin plays crucial roles in nearly every aspect of plant growth and development. Auxin signaling is activated through the phytohormone-induced proteasomal degradation of the Auxin/INDOLE-3-ACETIC ACID (Aux/IAA) family of transcriptional repressors. Notably, many auxin-modulated physiological processes are also regulated by nitric oxide (NO) that executes its biological effects predominantly through protein S-nitrosylation at specific cysteine residues. However, little is known about the molecular mechanisms in regulating the interactive NO and auxin networks. Here, we show that NO represses auxin signaling by inhibiting IAA17 protein degradation. NO induces the S-nitrosylation of Cys-70 located in the intrinsically disordered region of IAA17, which inhibits the TIR1-IAA17 interaction and consequently the proteasomal degradation of IAA17. The accumulation of a higher level of IAA17 attenuates auxin response. Moreover, an IAA17C70W nitrosomimetic mutation renders the accumulation of a higher level of the mutated protein, thereby causing partial resistance to auxin and defective lateral root development. Taken together, these results suggest that S-nitrosylation of IAA17 at Cys-70 inhibits its interaction with TIR1, thereby negatively regulating auxin signaling. This study provides unique molecular insights into the redox-based auxin signaling in regulating plant growth and development.

Early detection of lung cancer (LC) is vital for reducing LC-related mortality. However, noninvasive diagnostic tools remain a great challenge. We aim to identify blood-based biomarkers for the early detection of LC. Here, LC-associated hypomethylation in alpha-1,3-fucosyltransferase VII (FUT7) is identified via the Illumina 850K array in a discovery study and validated by mass spectrometry in two independent case-control studies with blood samples from 1720 LC patients (86.8% LC at stage I, blood is collected before surgery and treatment) and 3143 healthy controls. Compared to the controls, blood-based FUT7 hypomethylation is identified in LC patients at stage I, and even in LC patients with malignant nodules ≤ 1 cm and in patients with adenocarcinoma in situ. Gender plays a role in the LC-associated FUT7 hypomethylation in blood, which is more significant in males than in females. We also reveal that FUT7 hypomethylation in LC could be enhanced by the advanced stage of cancer, involvement of lymph nodes, and larger tumor size. Based on a large sample size and semi-quantitative methods, our study reveals a strong association between blood-based FUT7 hypomethylation and LC, suggesting that methylation signatures in blood may be a group of potential biomarkers for detection of early-stage LC.

The leading-edge CRISPR/CRISPR-associated technology is revolutionizing biotechnologies through genome editing. To track on/off-target events with emerging new editing techniques, improved bioinformatic tools are indispensable. Existing tools suffer from limitations in speed and scalability, especially with whole-genome sequencing (WGS) data analysis. To address these limitations, we have developed a comprehensive tool called CRISPR-detector, a web-based and locally deployable pipeline for genome editing sequence analysis. The core analysis module of CRISPR-detector is based on the Sentieon TNscope pipeline, with additional novel annotation and visualization modules designed to fit CRISPR applications. Co-analysis of the treated and control samples is performed to remove existing background variants prior to genome editing. CRISPR-detector offers optimized scalability, enabling WGS data analysis beyond Browser Extensible Data file-defined regions, with improved accuracy due to haplotype-based variant calling to handle sequencing errors. In addition, the tool also provides integrated structural variation calling and includes functional and clinical annotations of editing-induced mutations appreciated by users. These advantages facilitate rapid and efficient detection of mutations induced by genome editing events, especially for datasets generated from WGS. The web-based version of CRISPR-detector is available at https://db.cngb.org/crispr-detector, and the locally deployable version is available at https://github.com/hlcas/CRISPR-detector.

Olfactory receptors (ORs) play a key role in the prime sensorial perception, being highly relevant for intra/interspecific interactions. ORs are a subgroup of G-protein coupled receptors that exhibit highly complex subgenomes in vertebrates. However, OR repertoires remain poorly studied in fish lineages, precluding finely retracing their origin, evolution, and diversification, especially in the most basal groups. Here, we conduct an exhaustive gene screening upon 43 high-quality fish genomes exhibiting varied gene repertoires (2-583 genes). While the early vertebrates performed gas exchange through gills, we hypothesize that the emergence of new breathing structures (swim bladder and paired lungs) in early osteichthyans may be associated with expansions in the ORs gene families sensitive to airborne molecules. Additionally, we verify that the OR repertoire of moderns actinopterygians has not increased as expected following a whole genome duplication, likely due to regulatory mechanisms compensating the gene load excess. Finally, we identify 25 distinct OR families, allowing us to propose an updated universal nomenclature for the fish ORs.

As the only mammalian group capable of powered flight, bats have many unique biological traits. Previous comparative genomic studies in bats have focused on long-term evolution. However, the micro-evolutionary processes driving recent evolution are largely under-explored. Using resequencing data from 50 black flying foxes (Pteropus alecto), one of the model species for bats, we find that black flying fox has much higher genetic diversity and lower levels of linkage disequilibrium than most of the mammalian species. Demographic inference reveals strong population fluctuations (>100 fold) coinciding with multiple historical events including the last glacial change and Toba super eruption, suggesting that the black flying fox is a very resilient species with strong recovery abilities. While long-term adaptation in the black flying fox is enriched in metabolic genes, recent adaptation in the black flying fox has a unique landscape where recently selected genes are not strongly enriched in any functional category. The demographic history and mode of adaptation suggest that black flying fox might be a well-adapted species with strong evolutionary resilience. Taken together, this study unravels a vibrant landscape of recent evolution for the black flying fox and sheds light on several unique evolutionary processes for bats comparing to other mammalian groups.

MicroRNAs (miRNAs) are key regulators of myocyte development and traits, yet insight into the genetic basis of variation in miRNA expression is still limited. Here, we present a systematic analysis of expression quantitative trait loci (eQTL) for miRNA profiling in longissimus muscle of pigs from an eight-breed crossed heterogeneous population. By integrating the whole-genome sequencing and miRNAomics data, we map 54 cis- and 292 trans-eQTLs at high resolution that are associated with the expression of 54 and 92 miRNAs, respectively. Twenty-three trans-acting loci are identified to affect the expression of nine myomiRs (known muscle-specific miRNAs). MiRNAs in mammalian conserved miRNA clusters are found to be subjected to regulation by shared cis-eQTLs, while the expression of mature miRNA-5p/-3p counterparts is more likely to be regulated by different cis-eQTLs. Fine mapping and bioinformatics analyses pinpoint the peak cis-eSNP of miR-4331-5p, rs344650810, which is located in its seed region, as a causal variant for the changes in expression and function of this miRNA. Additionally, rs344650810 is significantly (P < 0.01) correlated with the density and percentage of type I muscle fibers. Altogether, this study provides a comprehensive atlas of miRNA-eQTLs in porcine skeletal muscle and new insights into regulatory mechanisms of miRNA expression.

Cytokinins influence many aspects of plant growth and development. Although cytokinin biosynthesis and signaling have been well studied in planta, little is known about the regulatory effects of epigenetic modifications on the cytokinin response. Here, we reveal that mutations to Morf Related Gene (MRG) proteins MRG1/MRG2, which are readers of trimethylated histone H3 lysine 4 and lysine 36 (H3K4me3 and H3K36me3), result in cytokinin hyposensitivity during various developmental processes, including callus induction and root and seedling growth inhibition. Similar to the mrg1 mrg2 mutant, plants with a defective AtTCP14, which belongs to the TEOSINTE BRANCHED, CYCLOIDEA, AND PROLIFERATING CELL FACTOR (TCP) transcription factor family, are insensitive to cytokinin. Furthermore, the transcription of several genes related to cytokinin signaling pathway is altered. Specifically, the expression of Arabidopsis thalianaHISTIDINE-CONTAINING PHOSPHOTRANSMITTER PROTEIN 2 (AHP2) decreases significantly in the mrg1 mrg2 and tcp14-2 mutants. We also confirm the interaction between MRG2 and TCP14 in vitro and in vivo. Thus, MRG2 and TCP14 can be recruited to AHP2 after recognizing H3K4me3/H3K36me3 markers and promote the histone-4 lysine-5 acetylation to further enhance AHP2 expression. In summary, our research elucidate a previously unknown mechanism mediating the effects of MRG proteins on the magnitude of the cytokinin response.