Zinc Finger Gene Research Articles

A loss-of-function mutation in OsTZF5 confers sensitivity to low temperature and effects the growth and development in rice.

Rice (Oryza sativa L.) is highly sensitive to low temperatures, which can significantly reduce its production. Cold tolerance in rice is a complex trait regulated by multiple mechanisms. OsTZF5, a member of the CCCH-type zinc finger gene family in rice, has been previously reported that overexpressing OsTZF5 under the stress-responsive promoter can confer drought resistance. In this study, we showed that the loss of function mutants of OsTZF5 decreased seed germination rate and chilling tolerance in rice, and influencing normal growth and development. OsTZF5 is expressed in various parts of the rice plant, including roots, stems, leaves and inflorescences, with the highest expression levels observed in leaves. Additionally, the expression of OsTZF5 gene was influenced by various stress conditions and hormone treatments. OsTZF5 knock-out mutants exhibited significantly lower survival rates compared to the wild type (Zhonghua11, ZH11) after cold stress, as well as fewer tillers, lower thousand-grain weight, and reduced grain yield under normal conditions. Transcriptomic analyses revealed that the expression of cold stress-related genes was significantly down-regulated in OsTZF5 knock-out mutants compared to ZH11 after cold stress. This down-regulation likely contributes to the reduced cold stress tolerance observed in OsTZF5 knock-out mutants. Our findings suggest that OsTZF5 is a multifunctional gene that plays a crucial role in regulating cold stress in rice.

Brassinosteroid signaling represses ZINC FINGER PROTEIN11 to regulate ovule development in Arabidopsis.

Brassinosteroid (BR) signaling and the C-class MADS-box gene AGAMOUS (AG) play important roles in ovule development in Arabidopsis (Arabidopsis thaliana). However, how BR signaling integrates with AG functions to control the female reproductive process remains elusive. Here, we showed that the regulatory role of BR signaling in proper ovule development is mediated by the transcriptional repressor gene ZINC FINGER PROTEIN 11 (ZFP11), which is a direct target of AG. ZFP11 expression initiates from the placenta upon AG induction and becomes prominent in the funiculus of ovule primordia. Plants harboring zfp11 mutations showed reduced placental length with decreased ovule numbers and some aborted ovules. During ovule development, the transcription factor BRASSINAZOLE-RESISTANT 1 (BZR1), which functions downstream of BR signaling, inhibits ZFP11 expression in the chalaza and nucellus. Weakened BR signaling leads to stunted integuments in ovules, resulting from the direct repression of INNER NO OUTER (INO) and WUSCHEL (WUS) by extended ZFP11 expression in the chalaza and nucellus, respectively. In addition, the zfp11 mutant shows reduced sensitivity to BR biosynthesis inhibitors and can rescue outer integument defects in brassinosteroid insensitive 1 (bri1) mutants. Thus, the precise spatial regulation of ZFP11, which is activated by AG in the placenta and suppressed by BR signaling in the central and distal regions of ovules, is essential for ensuring sufficient ovule numbers and proper ovule formation.

Genome-Wide Characterization of the INDETERMINATE DOMAIN (IDD) Zinc Finger Gene Family in Solanum lycopersicum and the Functional Analysis of SlIDD15 in Shoot Gravitropism.

The plant-specific IDD transcription factors (TFs) are vital for regulating plant growth and developmental processes. However, the characteristics and biological roles of the IDD gene family in tomato (Solanum lycopersicum) are still largely unexplored. In this study, 17 SlIDD genes were identified in the tomato genome and classified into seven subgroups according to the evolutionary relationships of IDD proteins. Analysis of exon-intron structures and conserved motifs reflected the evolutionary conservation of SlIDDs in tomato. Collinearity analysis revealed that segmental duplication promoted the expansion of the SlIDD family. Ka/Ks analysis indicated that SlIDD gene orthologs experienced predominantly purifying selection throughout evolution. The analysis of cis-acting elements revealed that the promoters of SlIDD genes contain numerous elements associated with light, plant hormones, and abiotic stresses. The RNA-seq data and qRT-PCR experimental results showed that the SlIDD genes exhibited tissue-specific expression. Additionally, Group A members from Arabidopsis thaliana and rice are known to play a role in regulating plant shoot gravitropism. QRT-PCR analysis confirmed that the expression level of SlIDD15 in Group A was high in the hypocotyls and stems. Subcellular localization demonstrated that the SlIDD15 protein was localized in the nucleus. Surprisingly, the loss-of-function of SlIDD15 by CRISPR/Cas9 gene editing technology did not display obvious gravitropic response defects, implying the existence of functional redundant factors within SlIDD15. Taken together, this study offers foundational insights into the tomato IDD gene family and serves as a valuable guide for exploring their molecular mechanisms in greater detail.

Hyd/UBR5 defines a tumor suppressor pathway that links Polycomb repressive complex to regulated protein degradation in tissue growth control and tumorigenesis.

Tumor suppressor genes play critical roles in normal tissue homeostasis, and their dysregulation underlies human diseases including cancer. Besides human genetics, model organisms such as Drosophila have been instrumental in discovering tumor suppressor pathways that were subsequently shown to be highly relevant in human cancer. Here we show that hyperplastic disc (Hyd), one of the first tumor suppressors isolated genetically in Drosophila and encoding an E3 ubiquitin ligase with hitherto unknown substrates, and Lines (Lin), best known for its role in embryonic segmentation, define an obligatory tumor suppressor protein complex (Hyd-Lin) that targets the zinc finger-containing oncoprotein Bowl for ubiquitin-mediated degradation, with Lin functioning as a substrate adaptor to recruit Bowl to Hyd for ubiquitination. Interestingly, the activity of the Hyd-Lin complex is directly inhibited by a micropeptide encoded by another zinc finger gene, drumstick (drm), which functions as a pseudosubstrate by displacing Bowl from the Hyd-Lin complex, thus stabilizing Bowl. We further identify the epigenetic regulator Polycomb repressive complex1 (PRC1) as a critical upstream regulator of the Hyd-Lin-Bowl pathway by directly repressing the transcription of the micropeptide drm Consistent with these molecular studies, we show that genetic inactivation of Hyd, Lin, or PRC1 resulted in Bowl-dependent hyperplastic tissue overgrowth in vivo. We also provide evidence that the mammalian homologs of Hyd (UBR5, known to be recurrently dysregulated in various human cancers), Lin (LINS1), and Bowl (OSR1/2) constitute an analogous protein degradation pathway in human cells, and that OSR2 promotes prostate cancer tumorigenesis. Altogether, these findings define a previously unrecognized tumor suppressor pathway that links epigenetic program to regulated protein degradation in tissue growth control and tumorigenesis.

Methylome analysis in girls with idiopathic central precocious puberty

BackgroundGenetic and environmental factors are implicated in many developmental processes. Recent evidence, however, has suggested that epigenetic changes may also influence the onset of puberty or the susceptibility to a wide range of diseases later in life. The present study aims to investigate changes in genomic DNA methylation profiles associated with pubertal onset analyzing human peripheral blood leukocytes from three different groups of subjects: 19 girls with central precocious puberty (CPP), 14 healthy prepubertal girls matched by age and 13 healthy pubertal girls matched by pubertal stage. For this purpose, the comparisons were performed between pre- and pubertal controls to identify changes in normal pubertal transition and CPP versus pre- and pubertal controls.ResultsAnalysis of methylation changes associated with normal pubertal transition identified 1006 differentially methylated CpG sites, 86% of them were found to be hypermethylated in prepubertal controls. Some of these CpG sites reside in genes associated with the age of menarche or transcription factors involved in the process of pubertal development. Analysis of methylome profiles in CPP patients showed 65% and 55% hypomethylated CpG sites compared with prepubertal and pubertal controls, respectively. In addition, interestingly, our results revealed the presence of 43 differentially methylated genes coding for zinc finger (ZNF) proteins. Gene ontology and IPA analysis performed in the three groups studied revealed significant enrichment of them in some pathways related to neuronal communication (semaphorin and gustation pathways), estrogens action, some cancers (particularly breast and ovarian) or metabolism (particularly sirtuin).ConclusionsThe different methylation profiles of girls with normal and precocious puberty indicate that regulation of the pubertal process in humans is associated with specific epigenetic changes. Differentially methylated genes include ZNF genes that may play a role in developmental control. In addition, our data highlight changes in the methylation status of genes involved in signaling pathways that determine the migration and function of GnRH neurons and the onset of metabolic and neoplastic diseases that may be associated with CPP in later life.

SETDB1 deletion causes DNA demethylation and upregulation of multiple zinc-finger genes

BackgroundSETDB1 (SET domain bifurcated-1) is a histone H3-lysine 9 (H3K9)-specific methyltransferase that mediates heterochromatin formation and repression of target genes. Despite the assumed functional link between DNA methylation and SETDB1-mediated H3K9 trimethylations, several studies have shown that SETDB1 operates autonomously of DNA methylation in a region- and cell-specific manner. This study analyzes SETDB1-null HAP1 cells through a linked methylome and transcriptome analysis, intending to explore genes controlled by SETDB1-involved DNA methylation.Methods and resultsWe investigated SETDB1-mediated regulation of DNA methylation and gene transcription in human HAP1 cells using reduced-representation bisulfite sequencing (RRBS) and RNA sequencing. While two-thirds of differentially methylated CpGs (DMCs) in genic regions were hypomethylated in SETDB1-null cells, we detected a plethora of C2H2-type zinc-finger protein genes (C2H2-ZFP, 223 of 749) among the DMC-associated genes. Most C2H2-ZFPs with DMCs in their promoters were found hypomethylated in SETDB1-KO cells, while other non-ZFP genes with promoter DMCs were not. These C2H2-ZFPs with DMCs in their promoters were significantly upregulated in SETDB1-KO cells. Similarly, C2H2-ZFP genes were upregulated in SETDB1-null 293T cells, suggesting that SETDB1’s function in ZFP gene repression is widespread. There are several C2H2-ZFP gene clusters on chromosome 19, which were selectively hypomethylated in SETDB1-KO cells.ConclusionsSETDB1 collectively and specifically represses a substantial fraction of the C2H2-ZFP gene family. Through the en-bloc silencing of a set of ZFP genes, SETDB1 may help establish a panel of ZFP proteins that are expressed cell-type specifically and thereby can serve as signature proteins for cellular identity.

Proteogenomic characterization of difficult-to-treat breast cancer with tumor cells enriched through laser microdissection

BackgroundBreast cancer (BC) is the most commonly diagnosed cancer and the leading cause of cancer death among women globally. Despite advances, there is considerable variation in clinical outcomes for patients with non-luminal A tumors, classified as difficult-to-treat breast cancers (DTBC). This study aims to delineate the proteogenomic landscape of DTBC tumors compared to luminal A (LumA) tumors.MethodsWe retrospectively collected a total of 117 untreated primary breast tumor specimens, focusing on DTBC subtypes. Breast tumors were processed by laser microdissection (LMD) to enrich tumor cells. DNA, RNA, and protein were simultaneously extracted from each tumor preparation, followed by whole genome sequencing, paired-end RNA sequencing, global proteomics and phosphoproteomics. Differential feature analysis, pathway analysis and survival analysis were performed to better understand DTBC and investigate biomarkers.ResultsWe observed distinct variations in gene mutations, structural variations, and chromosomal alterations between DTBC and LumA breast tumors. DTBC tumors predominantly had more mutations in TP53, PLXNB3, Zinc finger genes, and fewer mutations in SDC2, CDH1, PIK3CA, SVIL, and PTEN. Notably, Cytoband 1q21, which contains numerous cell proliferation-related genes, was significantly amplified in the DTBC tumors. LMD successfully minimized stromal components and increased RNA–protein concordance, as evidenced by stromal score comparisons and proteomic analysis. Distinct DTBC and LumA-enriched clusters were observed by proteomic and phosphoproteomic clustering analysis, some with survival differences. Phosphoproteomics identified two distinct phosphoproteomic profiles for high relapse-risk and low relapse-risk basal-like tumors, involving several genes known to be associated with breast cancer oncogenesis and progression, including KIAA1522, DCK, FOXO3, MYO9B, ARID1A, EPRS, ZC3HAV1, and RBM14. Lastly, an integrated pathway analysis of multi-omics data highlighted a robust enrichment of proliferation pathways in DTBC tumors.ConclusionsThis study provides an integrated proteogenomic characterization of DTBC vs LumA with tumor cells enriched through laser microdissection. We identified many common features of DTBC tumors and the phosphopeptides that could serve as potential biomarkers for high/low relapse-risk basal-like BC and possibly guide treatment selections.

Genome-wide analysis of C2H2 zinc finger gene family and its response to drought stress in common bean (Phaseolus vulgaris L.)

Genome-wide analysis of C2H2 zinc finger gene family and its response to drought stress in common bean (Phaseolus vulgaris L.)

Genome-wide identification, characterization and expression of C2H2 zinc finger gene family in Opisthopappus species under salt stress

BackgroundThe C2H2 zinc finger protein family plays important roles in plants. However, precisely how C2H2s function in Opisthopappus (Opisthopappus taihangensis and Opisthopappus longilobus) remains unclear.ResultsIn this study, a total of 69 OpC2H2 zinc finger protein genes were identified and clustered into five Groups. Seven tandem and ten fragment repeats were found in OpC2H2s, which underwent robust purifying selection. Of the identified motifs, motif 1 was present in all OpC2H2s and conserved at important binding sites. Most OpC2H2s possessed few introns and exons that could rapidly activate and react when faced with stress. The OpC2H2 promoter sequences mainly contained diverse regulatory elements, such as ARE, ABRE, and LTR. Under salt stress, two up-regulated OpC2H2s (OpC2H2-1 and OpC2H2-14) genes and one down-regulated OpC2H2 gene (OpC2H2-7) might serve as key transcription factors through the ABA and JA signaling pathways to regulate the growth and development of Opisthopappus species.ConclusionThe above results not only help to understand the function of C2H2 gene family but also drive progress in genetic improvement for the salt tolerance of Opisthopappus species.

The role and mechanism of the zinc finger gene ZNF580 in foam cell formation

Coronary atherosclerotic heart disease is an important threat to human health. The pathological basis is atherosclerosis, and foam cell formation is the key link in the initiation of atherosclerosis. Here, foam cell models were established using 50 ng/ml oxidized low-density lipoprotein (ox-LDL) to stimulate in vitro cultures of THP-1 cells for 72 h. The expression of ZNF580, a Cys2–His2 (C2H2) zinc finger protein containing 172 amino acids that was originally cloned by screening a human aortic cDNA library, was measured in foam cells, and its interaction with various regulatory factors during foam cell formation was investigated. Oil red O (ORO) staining was used to observe cell morphology and intracellular lipid levels. Lentivirus transfection was used to induce high ZNF580 expression (Ad-ZNF580) and low ZNF580 expression (Si-ZNF580) in THP-1 cells, and a fluorescent inverted microscope was used to observe the distribution of ZNF580 immunofluorescence to deduce the transfection rate. RNA and total protein were extracted, and the expression levels of ZNF580, cluster of differentiation 36 (CD36), peroxisome proliferator activated receptor-γ (PPAR-γ), ATP-binding cassette transporter A1 (ABCA1) and apolipoprotein E (ApoE) were measured by real-time quantitative PCR. The protein levels were examined by western blotting to evaluate the interaction between ZNF580 and associated regulatory factors. ZNF580 can significantly increase the expression levels of ApoE and ABCA1 and significantly decrease the expression levels of CD36 and PPAR-γ, suggesting that ZNF580-mediated inhibition of foam cell formation is associated with the PPAR-γ-CD36 signalling pathway. Based on these findings, ZNF580 might be a potential therapeutic candidate for the treatment of coronary atherosclerotic heart disease. Zhangzhongbai: ZNF580 inhibit foam cell formation

Abstract 4385: ZCCHC3 and Efp are prognostic factors for triple-negative breast cancer and potentially modulate cell division-related pathway

Abstract Triple-negative breast cancer (TNBC) is an aggressive subtype of breast cancer with poor prognosis and limited therapeutic strategies, lacking expression of estrogen receptor, progesterone receptor, and human epidermal growth factor receptor 2. We previously demonstrated that estrogen responsive finger protein (Efp)/TRIM25 functions as a ubiquitin ligase that degrades cell cycle checkpoint 14-3-3 sigma and promotes cell growth of ER-positive breast cancer [Urano et al., Nature 417:871-875, 2002]. In TNBC cells, we further showed that Efp regulates cell cycle-related gene expression and promotes cell proliferation [Sato et al., Biochem Biophys Res Commun 624:81-88, 2022]. Efp also exerts a distinct function to activate cellular RNA sensors such as RIG-I (retinoic acid inducible gene I), which plays a role in the innate immunity [Gack et al., Nature 446:916-920, 2007]. Another immune-related gene zinc finger CCHC domain-containing protein 3 (ZCCHC3) was recently suggested as Efp interactor, although its pathological relevance in breast cancer remains elusive. We here aimed to clarify the role of ZCCHC3 in TNBC. Immunohistochemical analysis of TNBC tissues from over 100 Japanese patients showed that a positive ZCCHC3 immunoreactivity (IR) was significantly associated with shorter disease-free survival (DFS) and positively correlated with Efp IR. Notably, patients with both ZCCHC3 and Efp IR positivity showed much shorter DFS than those with ZCCHC3 positivity alone. Functional analysis of ZCCHC3 by siRNA silencing demonstrated that ZCCHC3 promotes the proliferation, migration, and cell cycle progression of MDA-MB-231 TNBC cells. We established long-term culturable TNBC patient-derived cells (TNBC-PDC) using spheroid culture technique and revealed that the spheroid growth was suppressed by ZCCHC3-specific siRNA treatment. Moreover, ZCCHC3-specific siRNA injection suppressed in vivo tumor growth of MDA-MB-231 cells inoculated in immunodeficient mice. RNA sequencing analysis of ZCCHC3-silenced TNBC cells showed that cell division-related pathway was enriched among downregulated genes, including non-structural maintenance of chromosomes condensin I complex subunit H (NCAPH). NCAPH silencing inhibited the proliferation of TNBC-PDCs and MDA-MB-231 cells. Overall, ZCCHC3 and Efp are potential poor prognostic factors and can be applied to alternative therapeutic options for TNBC. Citation Format: Akihiro Fujimoto, Kazuhiro Ikeda, Toshihiko Takeiwa, Keiichi Kinowaki, Takuya Ogura, Hidetaka Kawabata, Akihiko Osaki, Kuniko Horie, Satoshi Inoue. ZCCHC3 and Efp are prognostic factors for triple-negative breast cancer and potentially modulate cell division-related pathway [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 4385.

A maternal-to-zygotic-transition gene block on the zebrafish sex chromosome.

Wild zebrafish (Danio rerio) have a ZZ/ZW chromosomal sex-determination system with the major sex locus on the right arm of chromosome-4 (Chr4R) near the largest heterochromatic block in the genome, suggesting that Chr4R transcriptomics might differ from the rest of the genome. To test this hypothesis, we conducted an RNA-seq analysis of adult ZW ovaries and ZZ testes in the Nadia strain and identified 4 regions of Chr4 with different gene expression profiles. Unique in the genome, protein-coding genes in a 41.7 Mb section (Region-2) were expressed in testis but silent in ovary. The AB lab strain, which lacks sex chromosomes, verified this result, showing that testis-biased gene expression in Region-2 depends on gonad biology, not on sex-determining mechanism. RNA-seq analyses in female and male brains and livers validated reduced transcripts from Region-2 in somatic cells, but without sex specificity. Region-2 corresponds to the heterochromatic portion of Chr4R and its content of genes and repetitive elements distinguishes it from the rest of the genome. Region-2 lacks protein-coding genes with human orthologs; has zinc finger genes expressed early in zygotic genome activation; has maternal 5S rRNA genes, maternal spliceosome genes, a concentration of tRNA genes, and a distinct set of repetitive elements. The colocalization of (1) genes silenced in ovaries but not in testes that are (2) expressed in embryos briefly at the onset of zygotic genome activation; (3) maternal-specific genes for translation machinery; (4) maternal-specific spliceosome components; and (5) adjacent genes encoding miR-430, which mediates maternal transcript degradation, suggest that this is a maternal-to-zygotic-transition gene regulatory block.

The HIV-1 reservoir landscape in persistent elite controllers and transient elite controllers.

BACKGROUNDPersistent controllers (PCs) maintain antiretroviral-free HIV-1 control indefinitely over time, while transient controllers (TCs) eventually lose virological control. It is essential to characterize the quality of the HIV reservoir in terms of these phenotypes in order to identify the factors that lead to HIV progression and to open new avenues toward an HIV cure.METHODSThe characterization of HIV-1 reservoir from peripheral blood mononuclear cells was performed using next-generation sequencing techniques, such as full-length individual and matched integration site proviral sequencing (FLIP-Seq; MIP-Seq).RESULTSPCs and TCs, before losing virological control, presented significantly lower total, intact, and defective proviruses compared with those of participants on antiretroviral therapy (ART). No differences were found in total and defective proviruses between PCs and TCs. However, intact provirus levels were lower in PCs compared with TCs; indeed the intact/defective HIV-DNA ratio was significantly higher in TCs. Clonally expanded intact proviruses were found only in PCs and located in centromeric satellite DNA or zinc-finger genes, both associated with heterochromatin features. In contrast, sampled intact proviruses were located in permissive genic euchromatic positions in TCs.CONCLUSIONSThese results suggest the need for, and can give guidance to, the design of future research to identify a distinct proviral landscape that may be associated with the persistent control of HIV-1 without ART.FUNDINGInstituto de Salud Carlos III (FI17/00186, FI19/00083, MV20/00057, PI18/01532, PI19/01127 and PI22/01796), Gilead Fellowships (GLD22/00147). NIH grants AI155171, AI116228, AI078799, HL134539, DA047034, MH134823, amfAR ARCHE and the Bill and Melinda Gates Foundation.

Genome-wide identification and expression analysis of the C2H2-zinc finger transcription factor gene family and screening of candidate genes involved in floral development in Coptis teeta Wall. (Ranunculaceae).

Background: C2H2-zinc finger transcription factors comprise one of the largest and most diverse gene superfamilies and are involved in the transcriptional regulation of flowering. Although a large number of C2H2 zinc-finger proteins (C2H2-ZFPs) have been well characterized in a number of model plant species, little is known about their expression and function in Coptis teeta. C. teeta displays two floral phenotypes (herkogamy phenotypes). It has been proposed that the C2H2-zinc finger transcription factor family may play a crucial role in the formation of floral development and herkogamy observed in C. teeta. As such, we performed a genome-wide analysis of the C2H2-ZFP gene family in C. teeta. Results: The complexity and diversity of C. teeta C2H2 zinc finger proteins were established by evaluation of their physicochemical properties, phylogenetic relationships, exon-intron structure, and conserved motifs. Chromosome localization showed that 95 members of the C2H2 zinc-finger genes were unevenly distributed across the nine chromosomes of C. teeta, and that these genes were replicated in tandem and segmentally and had undergone purifying selection. Analysis of cis-acting regulatory elements revealed a possible involvement of C2H2 zinc-finger proteins in the regulation of phytohormones. Transcriptome data was then used to compare the expression levels of these genes during the growth and development of the two floral phenotypes (F-type and M-type). These data demonstrate that in groups A and B, the expression levels of 23 genes were higher in F-type flowers, while 15 genes showed higher expressions in M-type flowers. qRT-PCR analysis further revealed that the relative expression was highly consistent with the transcriptome data. Conclusion: These data provide a solid basis for further in-depth studies of the C2H2 zinc finger transcription factor gene family in this species and provide preliminary information on which to base further research into the role of the C2H2 ZFPs gene family in floral development in C. teeta.

RNA-Seq-Based WGCNA and Association Analysis Reveal the Key Regulatory Module and Genes Responding to Salt Stress in Wheat Roots.

Soil salinization is the main abiotic stressor faced by crops. An improved understanding of the transcriptional response to salt stress in roots, the organ directly exposed to a high salinity environment, can inform breeding strategies to enhance tolerance and increase crop yield. Here, RNA-sequencing was performed on the roots of salt-tolerant wheat breeding line CH7034 at 0, 1, 6, 24, and 48 h after NaCl treatment. Based on transcriptome data, a weighted gene co-expression network analysis (WGCNA) was constructed, and five gene co-expression modules were obtained, of which the blue module was correlated with the time course of salt stress at 1 and 48 h. Two GO terms containing 249 differentially expressed genes (DEGs) related to osmotic stress response and salt-stress response were enriched in the blue module. These DEGs were subsequently used for association analysis with a set of wheat germplasm resources, and the results showed that four genes, namely a Walls Are Thin 1-related gene (TaWAT), an aquaporin gene (TaAQP), a glutathione S-transfer gene (TaGST), and a zinc finger gene (TaZFP), were associated with the root salt-tolerance phenotype. Using the four candidate genes as hub genes, a co-expression network was constructed with another 20 DEGs with edge weights greater than 0.6. The network showed that TaWAT and TaAQP were mainly co-expressed with fifteen interacting DEGs 1 h after salt treatment, while TaGST and TaZFP were mainly co-expressed with five interacting DEGs 48 h after salt treatment. This study provides key modules and candidate genes for understanding the salt-stress response mechanism in wheat roots.

Identification and expression profiling of flowering-related microRNAs and their targets reveals complex regulatory networks associated with bolting time in spinach

Bolting is one of the most important limiting traits in vegetable crops, like spinach, when spurt to flowering before harvesting. The evidence indicates bolting is controlled by various environmental factors including temperature, photoperiod, stresses, and genetic factors such as lncRNAs and miRNAs, but there is no information about miRNA-mediated gene expressions and their roles in flowering-time regulatory networks in spinach. Therefore, to gain more insights into the molecular mechanisms of flowering and bolting, we undertook a comprehensive analysis of miRNAs and their expression profiles in two distinct spinach accessions, namely Kashan (early bolting) and Viroflay (late bolting) by employing cutting-edge high-throughput sequencing technology. Our investigation identified 720 and 750 miRNAs in Kashan and Viroflay, respectively. Remarkably, amid the conserved miRNAs, a convergence of 15 families, exemplified by miR156, miR160, miR164, miR166, miR169, and miR399, renowned for their pivotal roles in flowering/bolting temporal control, was discerned across both accessions. Concerning differentially expressed miRNAs (DEMs), 64 target genes were identified, of which 40 genes were classified into seven flowering-related pathways including age, autonomous, circadian clock, hormone, integrator, photoperiodism, and vernalization. Moreover, novel crosstalks between miRNAs and various flowering-associated genes operating in flowering-related pathways, such as miR156 and squamosa promoter-binding proteins, and miR159 and GAMYB genes in the GA flowering pathway, were identified. Additionally, miR172 influenced MADS-box and AP2-like, while miR319 family, miR164, miR160, and miR169 played distinct roles in modulating key factors associated with flowering processes, including Flowering locus T, FLOWERING PROMOTING FACTOR 1, NAC domain, auxin response factor genes, zinc finger genes, and inorganic phosphate transporter genes, showcasing their intricate regulatory involvement in flowering and bolting pathways. The findings of this study will extend our knowledge of miRNA-mediated flowering modulation and can be used in breeding programs aimed at manipulating the spinach floral regulation system.

Hidden genomic diversity drives niche partitioning in a cosmopolitan eukaryotic picophytoplankton.

Marine eukaryotic phytoplankton are fundamental to the marine food web, yet the lack of reference genomes or just a single genome representing a taxon has led to an underestimation of their taxonomic, adaptive, and functional diversity. Here, we integrated strain isolation with metagenomic binning to recover genomes from the cosmopolitan picophytoplankton genus Bathycoccus, traditionally considered monospecific. Our recovery and analysis of 37 Bathycoccus genomes delineated their global genomic diversity and established four evolutionary clades (BI, BII, BIII, BIV). Our metagenomic abundance survey revealed well-differentiated ecological niches and distinct biogeographic distributions for each clade, predominantly shaped by temperature, salinity, and nutrient availability. Comparative genomics analyses further revealed clade-specific genomic traits that underpin niche adaptation and contribute to the global prevalence of Bathycoccus. Our findings underscore temperature as a major driver of genome diversification in this genus, with clade divergences coinciding with major paleoclimatic events that influenced their contemporary thermal niches. Moreover, the unique enrichment of C2H2 zinc finger and ankyrin repeat gene families in polar-adapted clades suggests previously unrecognized cold-adaptation mechanisms in marine eukaryotic phytoplankton. Our study offers a comprehensive genomic landscape of this crucial eukaryotic picophytoplankton, providing insights into their microdiversity and adaptive evolution in response to changing environments.

Genome-wide identification of the C2H2 zinc finger gene family and expression analysis under salt stress in sweetpotato.

The higher plant transcription factor C2H2 zinc finger protein (C2H2-ZFP) is essential for plant growth, development, and stress response. There are limited studies on C2H2-ZFP genes in sweetpotato, despite a substantial number of C2H2-ZFP genes having been systematically found in plants. In this work, 178 C2H2-ZFP genes were found in sweetpotato, distributed randomly on 15 chromosomes, and given new names according to where they were located. These members of the zinc finger gene family are separated into six branches, as shown by the phylogenetic tree. 24 tandem repeats of IbZFP genes and 46 fragment repeats were identified, and a homology study revealed that IbZFP genes linked more regions with wild relative species of sweetpotato as well as rhizome plants like potato and cassava. And we analyzed the expression patterns of IbZFP genes during the early development of sweetpotato storage roots (SRs) and salt stress using transcriptome data, and identified 44 IbZFP genes that exhibited differences in expression levels during the early expansion of sweetpotato SRs in different varieties, and 92 IbZFP genes that exhibited differences in expression levels under salt stress in salt tolerant and salt sensitive sweetpotato varieties. Additionally, we cloned six IbZFP genes in sweetpotato and analyzed their expression patterns in different tissues, their expression patterns under abiotic stress and hormone treatment, and subcellular localization. The results showed that the IbZFP genes had tissue specificity in sweetpotato and were induced to varying degrees by drought and salt stress. ABA and GA3 treatments also affected the expression of the IbZFP genes. We selected IbZFP105, which showed significant differences in expression levels under salt stress and ABA treatment, to be heterologously expressed in Arabidopsis thaliana. We found that IbZFP105 OE lines exhibited higher tolerance to salt stress and ABA stress. This indicates that IbZFP105 can enhance the salt tolerance of plants. These results systematically identified the evolution and expression patterns of members of the C2H2-ZFP gene family in sweetpotato, providing a theoretical basis for studying the role of IbZFP genes in the development of sweetpotato SRs and in resistance to stress.

The successive emergence ofERVL-MaLRs inprimates.

Although the ERVL-mammalian-apparent LTR retrotransposons (MaLRs) are the fourth largest family of transposable elements in the human genome, their evolutionary history and relationship have not been thoroughly studied. In this study, through RepeatMasker annotations of some representative species and construction of phylogenetic tree by sequence similarity, all primate-specific MaLR members are found to descend from MLT1A1 retrotransposon. Comparative genomic analysis, transposition-in-transposition inference, and sequence feature comparisons consistently show that each MaLR member evolved from its predecessor successively and had a limited activity period during primate evolution. Accordingly, a novel MaLR member was discovered as successor of MSTB1 in Tarsiiformes. At last, the identification of candidate precursor and intermediate THE1A elements provides further evidence for the previously proposed arms race model between ZNF430/ZNF100 and THE1B/THE1A. Taken together, this study sheds light on the evolutionary history of MaLRs and can serve as a foundation for future research on their interactions with zinc finger genes, gene regulation, and human health implications.

Abstract LB_B21: Extracellular RNA signatures of mutant KRAS-driven lung cancer

Abstract RAS genes are the most frequently mutated oncogenes in cancer, yet the effects of oncogenic RAS signaling on the noncoding transcriptome remain unclear. We analyzed the transcriptomes of human airway and bronchial epithelial cells transformed with mutant KRAS to define the landscape of KRAS-regulated noncoding RNAs. We find that oncogenic KRAS signaling upregulates noncoding transcripts throughout the genome, many of which arise from transposable elements (TEs). These TE RNAs exhibit differential expression and are regulated by KRAB zinc-finger (KZNF) genes, which are broadly downregulated in mutant KRAS cells and lung adenocarcinomas in vivo. Using both affinity- and nanofiltration-based EV isolation approaches, we show that specific TE RNAs, lncRNAs, and mRNAs, some of which are prognostic for lung adenocarcinoma (LUAD) patient survival, are preferentially released in extracellular vesicles (EVs). Moreover, we find that inhibition of oncogenic KRAS signaling broadly reprograms the noncoding transcriptome, as evidenced by a substantial increase in TE RNA secretion. Mutant KRAS inhibition also increases the abundance of secreted lncRNAs and retained intron-containing transcripts, while decreasing the mRNA content of EVs. Notably, oncogenic KRAS signaling is required for the secretion of mRNAs from a set of 20 genes that are significantly associated with unfavorable clinical outcomes in LUAD. Our results indicate that mutant KRAS remodels the repetitive noncoding transcriptome, demonstrating the broad scope of intracellular and extracellular RNAs regulated by this oncogenic signaling pathway. Our study also suggests that both coding and noncoding RNAs that are secreted in EVs may serve as mutant KRAS-specific signatures for diagnosing lung cancer via minimally invasive RNA liquid biopsy technology, which we are currently developing as a companion diagnostic for mutant KRAS-targeting therapies. Citation Format: Sree Lakshmi Velandi Maroli, Roman E Reggiardo, Reem Khojah, Daniel H Kim. Extracellular RNA signatures of mutant KRAS-driven lung cancer [abstract]. In: Proceedings of the AACR-NCI-EORTC Virtual International Conference on Molecular Targets and Cancer Therapeutics; 2023 Oct 11-15; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2023;22(12 Suppl):Abstract nr LB_B21.