CCCH-type Zinc Finger Protein Research Articles

Search for Expression Marker Genes That Reflect the Physiological Conditions of Blossom End Enlargement Occurrence in Cucumber.

Blossom end enlargement (BEE) is a postharvest deformation that may be related to the influx of photosynthetic assimilates before harvest. To elucidate the mechanism by which BEE occurs, expression marker genes that indicate the physiological condition of BEE-symptomatic fruit are necessary. First, we discovered that preharvest treatment with a synthetic cytokinin, N-(2-Chloro-4-pyridyl)-N'-phenylurea (CPPU), promoted fruit growth and suppressed BEE occurrence. This suggests that excessive assimilate influx is not a main cause of BEE occurrence. Subsequently, the expression levels of seven sugar-starvation marker genes, CsSEF1, AS, CsFDI1, CsPID, CsFUL1, CsETR1, and CsERF1B, were compared among symptomatic and asymptomatic fruits, combined with and without CPPU treatment. Only CsSEF1 showed a higher expression level in asymptomatic fruits than in symptomatic fruits, regardless of CPPU treatment. This was then tested using fruits stored via the modified-atmosphere packaging technique, which resulted in a lower occurrence of BEE, and the asymptomatic fruits showed a higher CsSEF1 expression level than symptomatic fruits, regardless of the packaging method. CsSEF1 codes a CCCH-type zinc finger protein, and an increase in the expression of CsSEF1 was correlated with a decrease in the fruit respiration rate. Thus, CsSEF1 may be usable as a BEE expression marker gene.

Multiomics Analyses Reveal MsC3H29 Positively Regulates Flavonoid Biosynthesis to Improve Drought Resistance of Autotetraploid Cultivated Alfalfa (Medicago sativa L.).

Alfalfa (Medicago sativa subsp. sativa), the "queen of forage," is the most important perennial legume, with high productivity and an excellent nutritional profile. Medicago sativa subsp. falcata is a subspecies of the alfalfa complex and exhibits better drought tolerance. However, drought stress significantly hampers their development and yield. The molecular mechanisms underlying the aboveground and underground tissues of sativa and falcata responding to drought stress remain obscure. Here, we performed a comprehensive comparative analysis of the physiological and transcriptomic responses of sativa and falcata under drought stress. The results showed that photosynthesis was inhibited, and antioxidant enzymes were activated under drought stress. MsC3H29, a CCCH-type zinc finger protein, was identified as a hub gene through weighted gene coexpression network analysis (WGCNA) and was significantly induced by drought in underground tissue. The MsC3H29 protein was localized in the nucleus. Overexpression (OE) of MsC3H29 can increase the primary root length and fresh weight of transgenic alfalfa hairy roots, while RNA interference (RNAi) decreases them under drought stress. The 2',7'-dichlorodihydrofluorescein diacetate (H2DCFDA) staining revealed that MsC3H29 promoted drought tolerance of alfalfa hairy roots through decreasing ROS accumulation. The targeted metabolome analysis showed that the overexpression of MsC3H29 resulted in higher levels of accumulation for flavonoid monomers, including vicenin, daidzein, apigenin, isorhamnetin, quercetin, and tricin, in transgenic alfalfa hairy roots before and after drought stress, while RNAi led to a reduction. Our study provided a key candidate gene for molecular breeding to improve drought resistance in alfalfa.

A nanopore-based cucumber genome assembly reveals structural variations at two QTLs controlling hypocotyl elongation.

The Xishuangbanna (XIS) cucumber (Cucumis sativus var. xishuangbannanesis) is a semiwild variety that has many distinct agronomic traits. Here, long reads generated by Nanopore sequencing technology helped assembling a high-quality genome (contig N50 = 8.7 Mb) of landrace XIS49. A total of 10,036 structural/sequence variations (SVs) were identified when comparing with Chinese Long (CL), and known SVs controlling spines, tubercles, and carpel number were confirmed in XIS49 genome. Two QTLs of hypocotyl elongation under low light, SH3.1 and SH6.1, were fine-mapped using introgression lines (donor parent, XIS49; recurrent parent, CL). SH3.1 encodes a red-light receptor Phytochrome B (PhyB, CsaV3_3G015190). A ∼4 kb region with large deletion and highly divergent regions (HDRs) were identified in the promoter of the PhyB gene in XIS49. Loss of function of this PhyB caused a super-long hypocotyl phenotype. SH6.1 encodes a CCCH-type zinc finger protein FRIGIDA-ESSENTIAL LIKE (FEL, CsaV3_6G050300). FEL negatively regulated hypocotyl elongation but it was transcriptionally suppressed by long terminal repeats retrotransposon insertion in CL cucumber. Mechanistically, FEL physically binds to the promoter of CONSTITUTIVE PHOTOMORPHOGENIC 1a (COP1a), regulating the expression of COP1a and the downstream hypocotyl elongation. These above results demonstrate the genetic mechanism of cucumber hypocotyl elongation under low light.

Characterization and Expression Analysis of HbC3H66: Implications for Transcriptional Regulation in Rubber Biosynthesis and Abiotic Stress Responses in Hevea brasiliensis

CCCH-type zinc-finger protein (ZnF_CCCH) family is one of the most important transcription factors (TFs) linked to various biotic and abiotic stressors and physiological and developmental processes in plants. This study aims to clone and characterise the latex gene expression patterns, evolution and characteristics of the Hevea brasiliensis ZnF_CCCH domain-containing protein 66 (HbC3H66) gene. These results showed that the open reading frame (ORF) of the HbC3H66 gene was 2,106 bp, encoding 701 aa and the calculated molecular weight of the encoded protein was 76.52 kDa. The N-terminal region of HbC3H66 contains 2 ANK repeats in, PfK_2/FBPase-2 and 2 type zinc finger motifs. Phylogenetic analysis showed that the C3H66 amino acid from Hevea and other plant C3H66 were clustered into 1 group and could be used for evolutionary analysis. Semi-quantitative RT-PCR (sqRT-PCR) revealed that the HbC3H66 mRNA was abundance in high-yielding Hevea clones (RRIT251 and RRIM600) and treatment with 2.5 % of ethephon (Eth) induced HbC3H66 mRNA expression in the latex of 15 years old Hevea trees. The HbC3H66 gene was induced by water deficit and 0.5 M NaCl in the latex of 3-month-old Hevea. In conclusion, the HbC3H66 protein may play a role in DNA-binding transcriptional regulation in NR pathways. Which would provide a theoretical basis for understanding the evolution and functions of the Hevea C3H66 gene in rubber biosynthesis. HIGHLIGHTS CCCH-type zinc-finger proteins (ZnF_CCCH) are essential transcription factors (TFs) for protein binding activity in plant growth and stress tolerance. Hevea brasiliensis zinc finger CCCH domain-containing protein 66 (HbC3H66) gene was clone and characterized. HbC3H66 contains 2 ANK repeat in, Pfk_2/FBPase-2 and 2 type zinc finger motifs. Semi-quantitative RT-PCR revealed that HbC3H66 gene was abundance in high-yielding Hevea clones (RRIT251 and RRIM600) and ethylene was induced HbC3H66 mRNA expression in the latex of Hevea The HbC3H66 gene was induced by water deficit and 0.5 M NaCl in the latex of seedling of Hevea trees. The HbC3H66 protein play a role in DNA-binding transcriptional regulation in NR pathway. GRAPHICAL ABSTRACT

Identification of CCCH-type zinc finger antiviral protein 1 (ZAP) gene from Pacific white shrimp (Penaeus vannamei): Characterization and expression analysis in response to viral infection.

Zinc-finger proteins (ZFPs) are a huge family that exert multiple roles in the cells. ZFPs could be divided into nine types based on the numbers and positions of conserved Cys and His residues, in which CCCH-type ZFP was one of the most widely studied types. CCCH-type zinc finger antiviral protein 1 (ZAP), a CCCH-type ZFP that can inhibit the replication of certain RNA viruses and DNA viruses by mediating degradation of viral RNA and repressing mRNA translation, plays significant roles in the host innate immune defenses against viral infections. Presently, there have been numerous reports investigating the antiviral ability of ZAP, while no data is available about ZAP gene in the species of shrimps or even crustaceans. In this study, a novel protein containing CCCH-type zinc finger motifs (ZnF-CCCH), CCCH-type zinc finger antiviral protein 1 (ZAP) gene, was identified from Pacific white shrimp (Penaeus vannamei) and its role in antiviral immunity was further investigated. Similar to mammalian ZAPs, in addition to ZnF-CCCH, PvZAP also possesses central WWE domains and C-terminal PARP domain. Phylogenetic analysis showed that PvZAP was close to that of the crustacean Pacific oyster, separating from the cluster of vertebrate ZAP proteins. Upon in vivo infection by IHHNV, gene expression of PvZAP was strongly up-regulated in the hepatopancreas and gills of both adult and juvenile shrimps, where adult individuals showed higher fold changes of up-regulation than in juvenile individuals. These results suggested that PvZAP might play an important role in the innate immune defense of Pacific white shrimp against IHHNV infection. This allows us to gain new insights into the immunological function of ZAP in the innate immunity of shrimp species and even crustaceans.

Expression of candidate marker genes of sugar starvation is upregulated in growth-suppressed parthenocarpic cucumber fruit. Novel gene markers for sugar starvation in growth-suppressed cucumber fruit.

To examine the physiological change in the growth suppression and abortion of parthenocarpic cucumber fruit, the expression of candidate marker genes of sugar starvation in relation to growth activity was examined. Fruits that failed to start exponential growth seemed to eventually abort. Hexose concentration of fruits was low in growth-suppressed fruit and increased in normally growing fruit consistent with the vacuolization. The correlation matrix indicated that the transcript levels of the genes, except CsaV3_6G046050 and CsaV3_5G032930, had a highly significant negative correlation with the relative growth rate in fruit length and had highly significant mutual positive correlations, suggesting that the asparagine synthetase gene, Cucumis sativus putative CCCH-type zinc finger protein CsSEF1, C. sativus BTB/POZ domain-containing protein At1g63850-like, CsaV3_3G000800, CsaV3_3G041280, and CsaV3_7G032930 are good markers of sugar starvation in cucumber fruit. The expression of candidate marker genes together with the hexose analysis strongly suggests that severe sugar starvation is occurring in growth-suppressed fruit.

Masculinizer gene controls sexual differentiation in Hyphantria cunea.

The Masculinizer gene, Masc, encodes a lepidopteran-specific novel CCCH-type zinc finger protein, which controls sex determination and dosage compensation in Bombyx mori. Considering the potential application of it in pest control, it is necessary to investigate the function of Masc gene in Hyphantria cunea, a globally invasive forest pest. In the present study, we identified and functionally characterized the Masc gene, HcMasc, in H. cunea. Sequence analysis revealed that HcMASC contained the conserved CCCH-type zinc finger domain, nuclear localization signal, and male determining domain, in which the last was confirmed to be required for its masculinization in BmN cell line. However, expression data showed that unlike male-biased expression in B. mori, HcMasc gene expresses in main all developmental stages or tissues in both sexes. Clustered regularly interspaced palindromic repeats (CRISPR) / CRISPR-associated protein 9-based disruption of the common exons 1 and 3 of the HcMasc gene resulted in imbalanced sex ratio and abnormal external genitalia of both sexes. Our results suggest that the HcMasc gene is required for both male and female sexual differentiation and dosage compensation in H. cunea and provide a foundation for developing better strategies to control this pest.

Genomic Identification of CCCH-Type Zinc Finger Protein Genes Reveals the Role of HuTZF3 in Tolerance of Heat and Salt Stress of Pitaya (Hylocereus polyrhizus)

Pitaya (Hylocereus polyrhizus) is cultivated in a broad ecological range, due to its tolerance to drought, heat, and poor soil. The zinc finger proteins regulate gene expression at the transcriptional and post-transcriptional levels, by interacting with DNA, RNA, and proteins, to play roles in plant growth and development, and stress response. Here, a total of 81 CCCH-type zinc finger protein genes were identified from the pitaya genome. Transcriptomic analysis showed that nine of them, including HuTZF3, responded to both salt and heat stress. RT-qPCR results showed that HuTZF3 is expressed in all tested organs of pitaya, with a high level in the roots and stems, and confirmed that expression of HuTZF3 is induced by salt and heat stress. Subcellular localization showed that HuTZF3 is targeted in the processing bodies (PBs) and stress granules (SGs). Heterologous expression of HuTZF3 could improve both salt and heat tolerance in Arabidopsis, reduce oxidative stress, and improve the activity of catalase and peroxidase. Therefore, HuTZF3 may be involved in post-transcriptional regulation via localizing to PBs and SGs, contributing to both salt and heat tolerance in pitaya.

The CCCH-Type Zinc-Finger Protein GhC3H20 Enhances Salt Stress Tolerance in Arabidopsis thaliana and Cotton through ABA Signal Transduction Pathway

The CCCH zinc-finger protein contains a typical C3H-type motif widely existing in plants, and it plays an important role in plant growth, development, and stress responses. In this study, a CCCH zinc-finger gene, GhC3H20, was isolated and thoroughly characterized to regulate salt stress in cotton and Arabidopsis. The expression of GhC3H20 was up-regulated under salt, drought, and ABA treatments. GUS activity was detected in the root, stem, leaves, and flowers of ProGhC3H20::GUS transgenic Arabidopsis. Compared with the control, the GUS activity of ProGhC3H20::GUS transgenic Arabidopsis seedlings under NaCl treatment was stronger. Through the genetic transformation of Arabidopsis, three transgenic lines of 35S-GhC3H20 were obtained. Under NaCl and mannitol treatments, the roots of the transgenic lines were significantly longer than those of the wild-type (WT) Arabidopsis. The leaves of the WT turned yellow and wilted under high-concentration salt treatment at the seedling stage, while the leaves of the transgenic Arabidopsis lines did not. Further investigation showed that compared with the WT, the content of catalase (CAT) in the leaves of the transgenic lines was significantly higher. Therefore, compared with the WT, overexpression of GhC3H20 enhanced the salt stress tolerance of transgenic Arabidopsis. A virus-induced gene silencing (VIGS) experiment showed that compared with the control, the leaves of pYL156-GhC3H20 plants were wilted and dehydrated. The content of chlorophyll in pYL156-GhC3H20 leaves was significantly lower than those of the control. Therefore, silencing of GhC3H20 reduced salt stress tolerance in cotton. Two interacting proteins (GhPP2CA and GhHAB1) of GhC3H20 have been identified through a yeast two-hybrid assay. The expression levels of PP2CA and HAB1 in transgenic Arabidopsis were higher than those in the WT, and pYL156-GhC3H20 had expression levels lower than those in the control. GhPP2CA and GhHAB1 are the key genes involved in the ABA signaling pathway. Taken together, our findings demonstrate that GhC3H20 may interact with GhPP2CA and GhHAB1 to participate in the ABA signaling pathway to enhance salt stress tolerance in cotton.

A tandem CCCH type zinc finger protein gene CpC3H3 from Chimonanthus praecox promotes flowering and enhances drought tolerance in Arabidopsis

BackgroundCCCH-type zinc finger proteins play important roles in plant development and biotic/abiotic stress responses. Wintersweet (Chimonanthus praecox) is a popular ornamental plant with strong resistance to various stresses, which is a good material for exploring gene resource for stress response. In this study, we isolated a CCCH type zinc finger protein gene CpC3H3 (MZ964860) from flower of wintersweet and performed functional analysis with a purpose of identifying gene resource for floral transition and stress tolerance.ResultsCpC3H3 was predicted a CCCH type zinc finger protein gene encoding a protein containing 446 amino acids with five conserved C-X8-C-X5-C-X3-H motifs. CpC3H3 was localized in the cell membrane but with a nuclear export signal at the N-terminal. Transcripts of CpC3H3 were significantly accumulated in flower buds at floral meristem formation stage, and were induced by polyethylene glycol. Overexpression of CpC3H3 promoted flowering, and enhanced drought tolerance in transgenic A. thaliana. CpC3H3 overexpression affects the expression level of genes involved in flower inducement and stress responses. Further comparative studies on physiological indices showed the contents of proline and soluble sugar, activity of peroxidase and the rates of electrolyte leakage were significantly increased and the content of malondialdehyde and osmotic potential was significantly reduced in transgenic A. thaliana under PEG stress.ConclusionOverall, CpC3H3 plays a role in flowering inducement and drought tolerance in transgenic A. thaliana. The CpC3H3 gene has the potential to be used to promote flowering and enhance drought tolerance in plants.

Chrysanthemum MAF2 regulates flowering by repressing gibberellin biosynthesis in response to low temperature.

Chrysanthemum (Chrysanthemum morifolium) is well known as a photoperiod-sensitive flowering plant. However, it has also evolved into a temperature-sensitive ecotype. Low temperature can promote the floral transition of the temperature-sensitive ecotype, but little is known about the underlying molecular mechanisms. Here, we identified MADS AFFECTING FLOWERING 2 (CmMAF2), a putative MADS-box gene, which induces floral transition in response to low temperatures independent of day length conditions in this ecotype. CmMAF2 was shown to bind to the promoter of the GA biosynthesis gene CmGA20ox1 and to directly regulate the biosynthesis of bioactive GA1 and GA4 . The elevated bioactive GA levels activated LEAFY (CmLFY) expression, ultimately initiating floral transition. In addition, CmMAF2 expression in response to low temperatures was directly activated by CmC3H1, a CCCH-type zinc-finger protein upstream. In summary, our results reveal that the CmC3H1-CmMAF2 module regulates flowering time in response to low temperatures by regulating GA biosynthesis in the temperature-sensitive chrysanthemum ecotype.

Transcriptome-Wide Identification of CCCH-Type Zinc Finger Proteins Family in Pinus massoniana and RR-TZF Proteins in Stress Response.

CCCH-type zinc finger proteins play an important role in multiple biotic and abiotic stresses. More and more reports about CCCH functions in plant development and stress responses have appeared over the past few years, focusing especially on tandem CCCH zinc finger proteins (TZFs). However, this has not been reported in Pinaceae. In this study, we identified 46 CCCH proteins, including 6 plant TZF members in Pinus massoniana, and performed bioinformatic analysis. According to RT-PCR analysis, we revealed the expression patterns of five RR-TZF genes under different abiotic stresses and hormone treatments. Meanwhile, tissue-specific expression analysis suggested that all genes were mainly expressed in needles. Additionally, RR-TZF genes showed transcriptional activation activity in yeast. The results in this study will be beneficial in improving the stress resistance of P. massoniana and facilitating further studies on the biological and molecular functions of CCCH zinc finger proteins.

AaZFP3, a Novel CCCH-Type Zinc Finger Protein from Adonis amurensis, Promotes Early Flowering in Arabidopsis by Regulating the Expression of Flowering-Related Genes.

CCCH-type zinc finger proteins (ZFP) are a large family of proteins that play various important roles in plant growth and development; however, the functions of most proteins in this family are uncharacterized. In this study, a CCCH-type ZFP, AaZFP3, was identified in the floral organ of Adonis amurensis. Quantitative real-time PCR (qPCR) analysis revealed that AaZFP3 was widely expressed in the flowers of A. amurensis. Subcellular localization analysis showed that the AaZFP3 protein was mainly localized to the cytoplasm in tobacco and Arabidopsis. Furthermore, the overexpression of AaZFP3 promoted early flowering in Arabidopsis under both normal and relatively low-temperature conditions. RNA-sequencing and qPCR analyses revealed that the expression of multiple key flowering-time genes was altered in transgenic Arabidopsis overexpressing AaZFP3 compared to wild-type. Of these genes, FLOWERING LOCUS T (AtFT) expression was most significantly up-regulated, whereas FLOWERING LOCUS C (AtFLC) was significantly down-regulated. These results suggest that the overexpression of AaZFP3 promotes early flowering in Arabidopsis by affecting the expression of flowering-time genes. Overall, our study indicates that AaZFP3 may be involved in flowering regulation in A. amurensis and may represent an important genetic resource for improving flowering-time control in other ornamental plants or crops.

Zinc Finger Protein ZFP36L1 Inhibits Flavivirus Infection by both 5'-3' XRN1 and 3'-5' RNA-Exosome RNA Decay Pathways.

Zinc-finger protein 36, CCCH type-like 1 (ZFP36L1), containing tandem CCCH-type zinc-finger motifs with an RNA-binding property, plays an important role in cellular RNA metabolism mainly by RNA decay pathways. Recently, we demonstrated that human ZFP36L1 has potent antiviral activity against influenza A virus infection. However, its role in the host defense response against flaviviruses has not been addressed. Here, we demonstrate that ZFP36L1 functions as a host innate defender against flaviviruses, including Japanese encephalitis virus (JEV) and dengue virus (DENV). Overexpression of ZFP36L1 reduced JEV and DENV infection, and ZFP36L1 knockdown enhanced viral replication. ZFP36L1 destabilized the JEV genome by targeting and degrading viral RNA mediated by both 5'-3' XRN1 and 3'-5' RNA-exosome RNA decay pathways. Mutation in both zinc-finger motifs of ZFP36L1 disrupted RNA-binding and antiviral activity. Furthermore, the viral RNA sequences specifically recognized by ZFP36L1 were mapped to the 3'-untranslated region of the JEV genome with the AU-rich element (AUUUA) motif. We extend the function of ZFP36L1 to host antiviral defense by directly binding and destabilizing the viral genome via recruiting cellular mRNA decay machineries. IMPORTANCE Cellular RNA-binding proteins are among the first lines of defense against various viruses, particularly RNA viruses. ZFP36L1 belongs to the CCCH-type zinc-finger protein family and has RNA-binding activity; it has been reported to bind directly to the AU-rich elements (AREs) of a subset of cellular mRNAs and then lead to mRNA decay by recruiting mRNA-degrading enzymes. However, the antiviral potential of ZFP36L1 against flaviviruses has not yet been fully demonstrated. Here, we reveal the antiviral potential of human ZFP36L1 against Japanese encephalitis virus (JEV) and dengue virus (DENV). ZFP36L1 specifically targeted the ARE motif within viral RNA and triggered the degradation of viral RNA transcripts via cellular degrading enzymes 5'-3' XRN1 and 3'-5' RNA exosome. These findings provide mechanistic insights into how human ZFP36L1 serves as a host antiviral factor to restrict flavivirus replication.

PIN and CCCH Zn-finger domains coordinate RNA targeting in ZC3H12 family endoribonucleases.

The CCCH-type zinc finger (ZnF) containing ZC3H12 ribonucleases are crucial in post-transcriptional immune homoeostasis with ZC3H12A being the only structurally studied member of the family. In this study, we present a structural-biochemical characterization of ZC3H12C, which is linked with chronic immune disorders like psoriasis. We established that the RNA substrate is cooperatively recognized by the PIN and ZnF domains of ZC3H12C and analyzed the crystal structure of ZC3H12C bound to a single-stranded RNA substrate. The RNA engages in hydrogen-bonded contacts and stacking interactions with the PIN and ZnF domains simultaneously. The ZC3H12 ZnF shows unprecedented structural features not previously observed in any member of the CCCH-ZnF family and utilizes stacking interactions via a unique combination of spatially conserved aromatic residues to align the target transcript in a bent conformation onto the ZnF scaffold. Further comparative structural analysis of ZC3H12 CCCH-ZnF suggests that a trinucleotide sequence is recognized by ZC3H12 ZnF in target RNA. Our work not only describes the initial structure-biochemical study on ZC3H12C, but also provides the first molecular insight into RNA recognition by a ZC3H12 family member. Finally, our work points to an evolutionary code for RNA recognition adopted by CCCH-type ZnF proteins.

DgC3H1, a CCCH zinc finger protein gene, confers cold tolerance in transgenic chrysanthemum

Zinc finger proteins play a significant role in regulating plant metabolism and stress responses. Nevertheless, the role of CCCH-type zinc finger proteins in chrysanthemums remains unclear. In this study, DgC3H1 was isolated and cloned from chrysanthemum. The DgC3H1 predicted protein contained 282 amino acids and three CCCH zinc finger motifs (C-X5-14-C-X4-5-C-X3-H). Through subcellular localization, it was found that DgC3H1 is located in the nucleus and is a CCCH zinc finger transcription factor. DgC3H1 overexpression improved the low temperature resistance of chrysanthemum. The expression of endogenous C3H1 was downregulated under antisense inhibition, and the sensitivity of chrysanthemum to low temperature stress was increased. Under cold stress, the contents of proline and soluble sugar, and the activities of peroxidase and superoxide dismutase were enhanced in the DgC3H1 overexpression lines,and reduced in the antisense suppression lines. In addition, the low temperature stress response genes DgCOR413, DgDREBa, DgCSD1, and DgCSD2 were upregulated in the overexpression lines, and downregulated in the antisense suppression lines when compared to the wild type. These results indicate that DgC3H1 can improve the resistance of chrysanthemum to cold stress and can be selected as an excellent candidate gene for breeding cold-tolerant plants.

ZC3H4-a novel Cys-Cys-Cys-His-type zinc finger protein-is essential for early embryogenesis in mice†.

Zinc finger domains of the Cys-Cys-Cys-His (CCCH) class are evolutionarily conserved proteins that bind nucleic acids and are involved in various biological processes. Nearly 60 CCCH-type zinc finger proteins have been identified in humans and mice, most have not been functionally characterized. Here, we provide the first in vivo functional characterization of ZC3H4-a novel CCCH-type zinc finger protein. Our results show that although Zc3h4 mutant embryos exhibit normal morphology at E3.5 blastocyst stage, they cannot be recovered at E7.5 early post-gastrulation stage, suggesting implantation failure. Outgrowth assays reveal that mutant blastocysts either fail to hatch from the zona pellucida, or can hatch but do not form a typical inner cell mass colony, the source of embryonic stem cells (ESCs). Although there is no change in levels of reactive oxygen species, Zc3h4 mutants display severe DNA breaks and reduced cell proliferation. Analysis of lineage specification reveals that both epiblast and primitive endoderm lineages are compromised with severe reductions in cell number and/or specification in the mutant blastocysts. In summary, these findings demonstrate the essential role of ZC3H4 during early mammalian embryogenesis.

Role of CCCH-Type Zinc Finger Proteins in Human Adenovirus Infections.

The zinc finger proteins make up a significant part of the proteome and perform a huge variety of functions in the cell. The CCCH-type zinc finger proteins have gained attention due to their unusual ability to interact with RNA and thereby control different steps of RNA metabolism. Since virus infections interfere with RNA metabolism, dynamic changes in the CCCH-type zinc finger proteins and virus replication are expected to happen. In the present review, we will discuss how three CCCH-type zinc finger proteins, ZC3H11A, MKRN1, and U2AF1, interfere with human adenovirus replication. We will summarize the functions of these three cellular proteins and focus on their potential pro- or anti-viral activities during a lytic human adenovirus infection.

Overexpression of Two CCCH-type Zinc-Finger Protein Genes Leads to Pollen Abortion in Brassica campestris ssp. chinensis.

The pollen grains produced by flowering plants are vital for sexual reproduction. Previous studies have shown that two CCCH-type zinc-finger protein genes in Brassica campestris, BcMF30a and BcMF30c, are involved in pollen development. Due to their possible functional redundancy, gain-of-function analysis is helpful to reveal their respective biological functions. Here, we found that the phenotypes of BcMF30a and BcMF30c overexpression transgenic plants driven by their native promoters were similar, suggesting their functional redundancy. The results showed that the vegetative growth was not affected in both transgenic plants, but male fertility was reduced. Further analysis found that the abortion of transgenic pollen was caused by the degradation of pollen contents from the late uninucleate microspore stage. Subcellular localization analysis demonstrated that BcMF30a and BcMF30c could localize in cytoplasmic foci. Combined with the studies of other CCCH-type genes, we speculated that the overexpression of these genes can induce the continuous assembly of abnormal cytoplasmic foci, thus resulting in defective plant growth and development, which, in this study, led to pollen abortion. Both the overexpression and knockout of BcMF30a and BcMF30c lead to abnormal pollen development, indicating that the appropriate expression levels of these two genes are critical for the maintenance of normal pollen development.

Zinc finger protein ZFP36L1 inhibits influenza A virus through translational repression by targeting HA, M and NS RNA transcripts.

ZFP36L1, a CCCH-type zinc finger protein, is an RNA-binding protein that participates in controlling cellular mRNA abundance and turnover by posttranscriptional regulation. Here, we demonstrated that ZFP36L1 has an important role in host defense against influenza A virus (IAV) infection. Overexpression of ZFP36L1 reduced IAV replication via translational repression of HA, M and NS RNA segment transcripts. IAV infection upregulated cellular ZFP36L1 expression, and endogenous ZFP36L1 knockdown significantly enhanced IAV replication. ZFP36L1 directly binds to IAV NS1 mRNA in the cytoplasm and blocks the expression and function of NS1 protein. Mutation of CCCH-type zinc finger domains of ZFP36L1 lost its antiviral potential and NS1 mRNA binding. Thus, ZFP36L1 can act as a host innate defense by targeting HA, M and NS mRNA transcripts to suppress viral protein translation.