Chromatin Assembly Factor Research Articles

Chromatin assembly factor 1B critically controls the early development but not function acquisition of invariant natural killer T cells in mice.

CD4+ CD8+ double-positive thymocytes give rise to both conventional TCRαβ+ T cells and invariant natural killer T cells (iNKT cells), but these two kinds of cells display different characteristics. The molecular mechanism underlying iNKT cell lineage development and function acquisition remain to be elucidated. We show that the loss of chromatin assembly factor 1B (CHAF1b) maintains the normal development of conventional TCRαβ+ T cells but severely impairs early development of iNKT cells. This dysregulation is accompanied by the impairment in chromatin activation and gene transcription at Vα14-Jα18 locus. Notably, ectopic expression of a Vα14-Jα18 TCR rescues Chaf1b-deficient iNKT cell developmental defects. Moreover, cytokine secretion and antitumor activity are substantially maintained in Vα14-Jα18 TCR transgene-rescued Chaf1b-deficient iNKT cells. Our study identifies CHAF1b as a critical factor that controls the early development but not function acquisition of iNKT cells via lineage- and stage-specific regulation.

De novo reconstitution of chromatin using wheat germ cell-free protein synthesis.

DNA is packaged with histones to form chromatin that impinges on all nuclear processes, including transcription, replication and repair, in the eukaryotic nucleus. A complete understanding of these molecular processes requires analysis of chromatin context in vitro. Here, Drosophila four core histones were produced in a native and unmodified form using wheat germ cell‐free protein synthesis. In the assembly reaction, four unpurified core histones and three chromatin assembly factors (dNAP‐1, dAcf1 and dISWI) were incubated with template DNA. We then assessed stoichiometry with the histones, nucleosome arrays, supercoiling and the ability of the chromatin to serve as a substrate for histone‐modifying enzymes. Overall, our method provides a new avenue to produce chromatin that can be useful in a wide range of chromatin research.

G2/M-checkpoint activation in fasciata1 rescues an aberrant S-phase checkpoint but causes genome instability.

The WEE1 and ATM AND RAD3-RELATED (ATR) kinases are important regulators of the plant intra-S-phase checkpoint; consequently, WEE1KO and ATRKO roots are hypersensitive to replication-inhibitory drugs. Here, we report on a loss-of-function mutant allele of the FASCIATA1 (FAS1) subunit of the chromatin assembly factor 1 (CAF-1) complex that suppresses the phenotype of WEE1- or ATR-deficient Arabidopsis (Arabidopsis thaliana) plants. We demonstrate that lack of FAS1 activity results in the activation of an ATAXIA TELANGIECTASIA MUTATED (ATM)- and SUPPRESSOR OF GAMMA-RESPONSE 1 (SOG1)-mediated G2/M-arrest that renders the ATR and WEE1 checkpoint regulators redundant. This ATM activation accounts for the telomere erosion and loss of ribosomal DNA that are described for fas1 plants. Knocking out SOG1 in the fas1 wee1 background restores replication stress sensitivity, demonstrating that SOG1 is an important secondary checkpoint regulator in plants that fail to activate the intra-S-phase checkpoint.

Histone chaperone CAF-1 promotes HIV-1 latency by leading the formation of phase-separated suppressive nuclear bodies.

HIV-1 latency is a major obstacle to achieving a functional cure for AIDS. Reactivation of HIV-1-infected cells followed by their elimination via immune surveillance is one proposed strategy for eradicating the viral reservoir. However, current latency-reversing agents (LRAs) show high toxicity and low efficiency, and new targets are needed to develop more promising LRAs. Here, we found that the histone chaperone CAF-1 (chromatin assembly factor 1) is enriched on the HIV-1 long terminal repeat (LTR) and forms nuclear bodies with liquid-liquid phase separation (LLPS) properties. CAF-1 recruits epigenetic modifiers and histone chaperones to the nuclear bodies to establish and maintain HIV-1 latency in different latency models and primary CD4+ T cells. Three disordered regions of the CHAF1A subunit are important for phase-separated CAF-1 nuclear body formation and play a key role in maintaining HIV-1 latency. Disruption of phase-separated CAF-1 bodies could be a potential strategy to reactivate latent HIV-1.

MiR-1179 is downregulated in cervical cancer and its overexpression suppresses cancer cells invasion by targeting CHAF1A/ZEB1.

The anticancer effect of miR-1179 has been extensively studied in many tumors. The mechanism of miR-1179 action in cervical cancer, however, remains largely unknown. In the present study, miR-1179 was downregulated in both cervical cancer cell lines and cancer tissues. In addition, miR-1179 mimic suppressed cancer cells invasion and epithelial-mesenchymal transition (EMT) in cervical cancer SiHa and Caski cells. We found that chromatin assembly factor 1 subunit A (CHAF1A) might be a direct target of miR-1179 and could be regulated by miR-1179. Furthermore, CHAF1A shRNA suppressed the cervical cancer cells invasion and the expression of EMT-promoted proteins. Reversely, CHAF1A overexpression not only promoted cervical cancer cells invasion but also upregulated the level of Zinc finger E-box bindingprotein1 (ZEB1), an EMT-related protein. The induction of ZEB1 could be counteracted by miR-1179 overexpression. It was observed that in cervical cancer patients' tissues, miR-1179 was downregulated while the pathway of CHAF1A/ZEB1 was upregulated. In summary, our research indicated that the miR-1179 might regulate CHAF1A/ZEB1 axis and inhibit the invasion of cervical cancer cells.

Five candidate biomarkers associated with the diagnosis and prognosis of cervical cancer.

Purpose: Cervical cancer (CC) is one of the most general gynecological malignancies and is associated with high morbidity and mortality. We aimed to select candidate genes related to the diagnosis and prognosis of CC.Methods: The mRNA expression profile datasets were downloaded. We also downloaded RNA-sequencing gene expression data and related clinical materials from TCGA, which included 307 CC samples and 3 normal samples. Differentially expressed genes (DEGs) were obtained by R software. GO function analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis of DEGs were performed in the DAVID dataset. Using machine learning, the optimal diagnostic mRNA biomarkers for CC were identified. We used qRT-PCR and Human Protein Atlas (HPA) database to exhibit the differences in gene and protein levels of candidate genes.Results: A total of 313 DEGs were screened from the microarray expression profile datasets. DNA methyltransferase 1 (DNMT1), Chromatin Assembly Factor 1, subunit B (CHAF1B), Chromatin Assembly Factor 1, subunit A (CHAF1A), MCM2, CDKN2A were identified as optimal diagnostic mRNA biomarkers for CC. Additionally, the GEPIA database showed that the DNMT1, CHAF1B, CHAF1A, MCM2 and CDKN2A were associated with the poor survival of CC patients. HPA database and qRT-PCR confirmed that these genes were highly expressed in CC tissues.Conclusion: The present study identified five DEmRNAs, including DNMT1, CHAF1B, CHAF1A, MCM2 and Kinetochore-related protein 1 (KNTC1), as potential diagnostic and prognostic biomarkers of CC.

Disruption of NAP1 genes in Arabidopsis thaliana suppresses the fas1 mutant phenotype, enhances genome stability and changes chromatin compaction.

Histone chaperones mediate the assembly and disassembly of nucleosomes and participate in essentially all DNA-dependent cellular processes. In Arabidopsis thaliana, loss-of-function of FAS1 or FAS2 subunits of the H3-H4 histone chaperone complex CHROMATIN ASSEMBLY FACTOR 1 (CAF-1) has a dramatic effect on plant morphology, growth and overall fitness. CAF-1 dysfunction can lead to altered chromatin compaction, systematic loss of repetitive elements or increased DNA damage, clearly demonstrating its severity. How chromatin composition is maintained without functional CAF-1 remains elusive. Here we show that disruption of the H2A-H2B histone chaperone NUCLEOSOME ASSEMBLY PROTEIN 1 (NAP1) suppresses the FAS1 loss-of-function phenotype. The quadruple mutant fas1 nap1;1 nap1;2 nap1;3 shows wild-type growth, decreased sensitivity to genotoxic stress and suppression of telomere and 45S rDNA loss. Chromatin of fas1 nap1;1 nap1;2 nap1;3 plants is less accessible to micrococcal nuclease and the nuclear H3.1 and H3.3 histone pools change compared to fas1. Consistently, association between NAP1 and H3 occurs in the cytoplasm and nucleus in vivo in protoplasts. Altogether we show that NAP1 proteins play an essential role in DNA repair in fas1, which is coupled to nucleosome assembly through modulation of H3 levels in the nucleus.

Chromatin assembly factor 1 suppresses epigenetic reprogramming toward adaptive drug resistance

The long-term effectiveness of targeted cancer therapies is limited by the development of resistance. Although epigenetic reprogramming has been implicated in resistance, the mechanisms remain elusive. Herein, we demonstrate that increased chromatin accessibility is involved in adaptive BRAF inhibitor (BRAFi)-resistance in melanoma cells. We observed loss of chromatin assembly factor 1 (CAF-1) and its related histone H3 lysine 9 trimethylation (H3K9me3) with adaptive BRAFi resistance. We further showed that depletion of CAF-1 provides chromatin plasticity for effective reprogramming by AP1 components to promote BRAFi resistance. Our data suggest that therapeutic approaches to restore H3K9me3 levels may compensate for the loss of CAF-1 and, in turn, suppress resistance to BRAF inhibitors.

Silencing Anti-Silencing Function 1B Inhibited Proliferation and Migration of Cervical Cancer Cells and Promoted Apoptosis by Down-Regulating Chromatin Assembly Factor 1, Subunit B

The expression of anti-silencing function 1B (ASF1B) in cervical cancer patients were previously reported, indicating its implications as a new candidate gene related to the development of cervical lesions and cancer. In this study, we investigated the possible role of ASF1B in cervical carcinogenesis. ASF1B messenger RNA and protein expression was assessed in five cervical cancer cell line models, Hela, C-33A, SiHa, CaSKi, HCC-94 and normal cervical epithelium cell Ect. Gene silencing approach was employed to investigate the potential role of ASF1B in cellular growth, proliferation, colony-forming ability, migration and invasion in Hela cells. Our data indicated that ASF1B was expressed in all cervical cancer cells at the gene and protein level. Gene silencing of ASF1B caused significant inhibition in cellular proliferation, colony-forming ability, migration and invasion ability, and promote apoptosis of Hela cells. However, the biological effects of ASF1B silencing on Hela cells were reversing after down-regulating recombinant chromatin assembly factor 1, subunit B (CHAF1B). Collectively, our findings concluded that silencing ASF1B inhibits proliferation and migration of Hela cells and promotes apoptosis by down-regulating CHAF1B.

CAF-1 and Rtt101p function within the replication-coupled chromatin assembly network to promote H4 K16ac, preventing ectopic silencing.

Replication-coupled chromatin assembly is achieved by a network of alternate pathways containing different chromatin assembly factors and histone-modifying enzymes that coordinate deposition of nucleosomes at the replication fork. Here we describe the organization of a CAF-1-dependent pathway in Saccharomyces cerevisiae that regulates acetylation of histone H4 K16. We demonstrate factors that function in this CAF-1-dependent pathway are important for preventing establishment of silenced states at inappropriate genomic sites using a crippled HMR locus as a model, while factors specific to other assembly pathways do not. This CAF-1-dependent pathway required the cullin Rtt101p, but was functionally distinct from an alternate pathway involving Rtt101p-dependent ubiquitination of histone H3 and the chromatin assembly factor Rtt106p. A major implication from this work is that cells have the inherent ability to create different chromatin modification patterns during DNA replication via differential processing and deposition of histones by distinct chromatin assembly pathways within the network.

Reconstitution of Drosophila and human chromatins by wheat germ cell-free co-expression system

BackgroundElaboration of the epigenetic regulation of chromatin is a long-standing aim in molecular and cellular biology. Hence, there is a great demand for the development of in vitro methods to reconstitute chromatin that can be used directly for biochemical assays. The widely used wheat germ cell-free protein expression method provides broad applications to investigate the function and structure of eukaryotic proteins. Such advantages, including high translation efficiency, flexibility, and possible automatization, are beneficial for achieving native-like chromatin substrates for in vitro studies.ResultsWe describe a novel, single-step in vitro chromatin assembly method by using the wheat germ cell-free protein synthesis. We demonstrated that both Drosophila and human chromatins can be reconstituted in the course of the in vitro translation of core histones by the addition of chromatin assembly factors, circular plasmid, and topoisomerase I in an ATP-dependent manner. Drosophila chromatin assembly was performed in 4 h at 26 °C, in the presence of premixed mRNAs encoding the core histones, dAcf1/dISWI chromatin remodeling complex, and nucleosome assembly protein, dNAP1. Similarly, the human chromatin was assembled by co-expressing the human core histones with Drosophila chromatin remodeling factor, dISWI, and chromatin chaperone, dNLP, for 6 h at 26 °C. The presence of reconstituted chromatin was monitored by DNA supercoiling assay, also the regular spacing of nucleosomes was assessed by Micrococcal nuclease assay. Furthermore, Drosophila linker histone H1-containing chromatin was reconstituted, affirming that the in vitro assembled chromatin is suitable for downstream applications.ConclusionsThe method described in this study allows the assembly of Drosophila and human chromatins, possibly in native-like form, by using a wheat germ cell-free protein expression. Although both chromatins were reconstituted successfully, there were unexpected differences with respect to the required ratio of histone-coding mRNAs and the reaction time. Overall, our new in vitro chromatin reconstitution method will aid to characterize the unrevealed structure, function, and regulation of chromatin dynamics.

Reprogramming of the histone H3.3 landscape in the early mouse embryo.

Epigenetic reprogramming of the zygote involves dynamic incorporation of histone variant H3.3. However, the genome-wide distribution and dynamics of H3.3 during early development remain unknown. Here, we delineate the H3.3 landscapes in mouse oocytes and early embryos. We unexpectedly identify a non-canonical H3.3 pattern in mature oocytes and zygotes, in which local enrichment of H3.3 at active chromatin is suppressed and H3.3 is relatively evenly distributed across the genome. Interestingly, although the non-canonical H3.3 pattern forms gradually during oogenesis, it quickly switches to a canonical pattern at the two-cell stage in a transcription-independent and replication-dependent manner. We find that incorporation of H3.1/H3.2 mediated by chromatin assembly factor CAF-1 is a key process for the de novo establishment of the canonical pattern. Our data suggest that the presence of the non-canonical pattern and its timely transition toward a canonical pattern support the developmental program of early embryos.

Regulation of ALT-associated homology-directed repair by polyADP-ribosylation.

The synthesis of poly(ADP-ribose) (PAR) reconfigures the local chromatin environment and recruits DNA-repair complexes to damaged chromatin. PAR degradation by poly(ADP-ribose) glycohydrolase (PARG) is essential for progression and completion of DNA repair. Here, we show that inhibition of PARG disrupts homology-directed repair (HDR) mechanisms that underpin alternative lengthening of telomeres (ALT). Proteomic analyses uncover a new role for poly(ADP-ribosyl)ation (PARylation) in regulating the chromatin-assembly factor HIRA in ALT cancer cells. We show that HIRA is enriched at telomeres during the G2 phase and is required for histone H3.3 deposition and telomere DNA synthesis. Depletion of HIRA elicits systemic death of ALT cancer cells that is mitigated by re-expression of ATRX, a protein that is frequently inactivated in ALT tumors. We propose that PARylation enables HIRA to fulfill its essential role in the adaptive response to ATRX deficiency that pervades ALT cancers.

Large tandem duplications affect gene expression, 3D organization, and plant-pathogen response.

Rapid plant genome evolution is crucial to adapt to environmental changes. Chromosomal rearrangements and gene copy number variation (CNV) are two important tools for genome evolution and sources for the creation of new genes. However, their emergence takes many generations. In this study, we show that in Arabidopsis thaliana, a significant loss of ribosomal RNA (rRNA) genes with a past history of a mutation for the chromatin assembly factor 1 (CAF1) complex causes rapid changes in the genome structure. Using long-read sequencing and microscopic approaches, we have identified up to 15 independent large tandem duplications in direct orientation (TDDOs) ranging from 60 kb to 1.44 Mb. Our data suggest that these TDDOs appeared within a few generations, leading to the duplication of hundreds of genes. By subsequently focusing on a line only containing 20% of rRNA gene copies (20rDNA line), we investigated the impact of TDDOs on 3D genome organization, gene expression, and cytosine methylation. We found that duplicated genes often accumulate more transcripts. Among them, several are involved in plant–pathogen response, which could explain why the 20rDNA line is hyper-resistant to both bacterial and nematode infections. Finally, we show that the TDDOs create gene fusions and/or truncations and discuss their potential implications for the evolution of plant genomes.

CAF-1/p150 promotes cell proliferation, migration, invasion and predicts a poor prognosis in patients with cervical cancer.

Cervical cancer is one of the most common malignancies among women worldwide that exhibits high morbidity and mortality rates. Thus, the discovery of novel molecules and targets for cervical cancer diagnosis and treatment is critical. The present study aimed to investigate the role of the chromatin assembly factor (CAF)-1 subunit, CAF-1/p150 on cervical cancer cell proliferation, migration and invasion. Immunohistochemical analysis was used to detect the CAF-1/p150 expression in cervical cancer tissues and to analyze the association between CAF-1/p150 expression and the prognosis of patients with cervical cancer. In addition, colony formation, wound healing and Transwell assays were used to assess the function of CAF-1/p150 in cervical cancer cells. The results demonstrated that CAF-1/p150 was expressed in both normal and cervical cancer tissues. CAF-1/p150 protein expression was localized in the cell nuclei and was highly expressed in cervical cancer tissues. Furthermore, high CAF-1/p150 expression was significantly associated with FIGO stage, local recurrence, distant metastasis and a shorter overall survival time of patients with cervical cancer. CAF-1/p150 knockdown attenuated the anchorage-independent proliferation, migration and invasion of Hela and SiHa cervical cancer cells in vitro. Taken together, the results of the present study confirmed the involvement of CAF-1/p150 in the progression of cervical cancer, and validated its use as a poor prognostic indicator in patients with cervical cancer.

Ubiquitin ligase CHAF1B induces cisplatin resistance in lung adenocarcinoma by promoting NCOR2 degradation

BackgroundLung cancer is the most common malignant tumor in the world. The Whole-proteome microarray showed that ubiquitin ligase chromatin assembly factor 1 subunit B (CHAF1B) expression in A549/DDP cells is higher than in A549 cells. Our study explored the molecular mechanism of CHAF1B affecting cisplatin resistance in lung adenocarcinoma (LUAD).MethodsProteome microarray quantify the differentially expressed proteins between LUAD cell line A549 and its cisplatin-resistant strain A549/DDP. Quantitative real-time quantitative polymerase chain reaction (qRT-PCR) and Western blot (WB) confirmed the CHAF1B expression. Public databases analyzed the prognosis of LUAD patients with varied LUAD expression followed by the substrates prediction of CHAF1B. Public databases showed that nuclear receptor corepressor 2 (NCOR2) may be substrates of CHAF1B. WB detected that CHAF1B expression affected the expression of NCOR2. Cell and animal experiments and clinical data detected function and integrating mechanism of CHAF1B compounds.ResultsProteome chips results indicated that CHAF1B, PPP1R13L, and CDC20 was higher than A549 in A549/DDP. Public databases showed that high expression of CHAF1B, PPP1R13L, and CDC20 was negatively correlated with prognosis in LUAD patients. PCR and WB results indicated higher CHAF1B expression in A549/DDP cells than that in A549 cells. NCOR2 and PPP5C were confirmed to be substrates of CHAF1B. CHAF1B knockdown significantly increased the sensitivity of cisplatin in A549/DDP cells and the upregulated NCOR2 expression. CHAF1B and NCOR2 are interacting proteins and the position of interaction between CHAF1B and NCOR2 was mainly in the nucleus. CHAF1B promotes ubiquitination degradation of NCOR2. Cells and animal experiments showed that under the action of cisplatin, after knockdown of CHAF1B and NCOR2 in A549/DDP group compared with CHAF1B knockdown alone, the cell proliferation and migratory ability increased and apoptotic rate decreased, and the growth rate and size of transplanted tumor increased significantly. Immunohistochemistry suggested that Ki-67 increased, while apoptosis-related indicators caspase-3 decreased significantly. Clinical data showed that patients with high expression of CHAF1B are more susceptible to cisplatin resistance.ConclusionUbiquitin ligase CAHF1B can induce cisplatin resistance in LUAD by promoting the ubiquitination degradation of NCOR2.

The multifaceted histone chaperone RbAp46/48 in Plasmodium falciparum: structural insights, production, and characterization.

RbAp46/RBBP7 and RbAp48/RBBP4 are WD40-repeat histone chaperones and chromatin adaptors that reside in multiple complexes involved in maintenance of chromatin structure. RbAp48 is the essential subunit of the chromatin assembly factor-1 (CAF-1) complex, therefore also named as CAF-1C. A detailed in silico sequence and structure analysis of homologs of RbAp46/48 in Plasmodium falciparum (PF3D7_0110700 and PF3D7_1433300) exhibited conservation of characteristic features in both the protein-seven-bladed WD40 β-propeller conformation and different binding interfaces. A comparative structural analysis highlighted species-specific features of the parasite, yeast, drosophila, and human RbAp46/48. In the present study, we report cloning, expression, and characterization of P. falciparum PF3D7_0110700, a putative RbAp46/48 (PfRbAp46/48). PfRbAp46/48 was cloned into pTEM11 vector in fusion with 6xHistidine tag and over-expressed in Escherichia coli B834 cells. The protein was purified by Ni-NTA followed by gel permeation chromatography. The protein expressed in all the three asexual blood stages and exhibited nuclear localization. We showed direct interaction of the purified rPfRbAp46/48 with the histone H4. These findings further our understanding of RbAp46/48 proteins and role of these proteins in the parasite biology.

Deposition of Centromeric Histone H3 Variant CENP-A/Cse4 into Chromatin Is Facilitated by Its C-Terminal Sumoylation.

Centromeric localization of CENP-A (Cse4 in Saccharomyces cerevisiae, CID in flies, CENP-A in humans) is essential for faithful chromosome segregation. Mislocalization of overexpressed CENP-A contributes to aneuploidy in yeast, flies, and humans, and is proposed to promote tumorigenesis in human cancers. Hence, defining molecular mechanisms that promote or prevent mislocalization of CENP-A is an area of active investigation. In budding yeast, evolutionarily conserved histone chaperones Scm3 and chromatin assembly factor-1 (CAF-1) promote localization of Cse4 to centromeric and noncentromeric regions, respectively. Ubiquitin ligases, such as Psh1 and Slx5, and histone chaperones (HIR complex) regulate proteolysis of overexpressed Cse4 and prevent its mislocalization to noncentromeric regions. In this study, we have identified sumoylation sites lysine (K) 215/216 in the C terminus of Cse4, and shown that sumoylation of Cse4 K215/216 facilitates its genome-wide deposition into chromatin when overexpressed. Our results showed reduced levels of sumoylation of mutant Cse4 K215/216R/A [K changed to arginine (R) or alanine (A)] and reduced interaction of mutant Cse4 K215/216R/A with Scm3 and CAF-1 when compared to wild-type Cse4 Consistent with these results, levels of Cse4 K215/216R/A in the chromatin fraction and localization to centromeric and noncentromeric regions were reduced. Furthermore, in contrast to GAL-CSE4, which exhibits Synthetic Dosage Lethality (SDL) in psh1∆, slx5∆, and hir2∆ strains, GAL-cse4K215/216R does not exhibit SDL in these strains. Taken together, our results show that deposition of Cse4 into chromatin is facilitated by its C-terminal sumoylation.

Alu retrotransposons modulate Nanog expression through dynamic changes in regional chromatin conformation via aryl hydrocarbon receptor

Transcriptional repression of Nanog is an important hallmark of stem cell differentiation. Chromatin modifications have been linked to the epigenetic profile of the Nanog gene, but whether chromatin organization actually plays a causal role in Nanog regulation is still unclear. Here, we report that the formation of a chromatin loop in the Nanog locus is concomitant to its transcriptional downregulation during human NTERA-2 cell differentiation. We found that two Alu elements flanking the Nanog gene were bound by the aryl hydrocarbon receptor (AhR) and the insulator protein CTCF during cell differentiation. Such binding altered the profile of repressive histone modifications near Nanog likely leading to gene insulation through the formation of a chromatin loop between the two Alu elements. Using a dCAS9-guided proteomic screening, we found that interaction of the histone methyltransferase PRMT1 and the chromatin assembly factor CHAF1B with the Alu elements flanking Nanog was required for chromatin loop formation and Nanog repression. Therefore, our results uncover a chromatin-driven, retrotransposon-regulated mechanism for the control of Nanog expression during cell differentiation.

Unraveling CAF-1 family in Plasmodium falciparum: comparative genome-wide identification and phylogenetic analysis among eukaryotes, expression profiling and protein-protein interaction studies.

The present research reports a detailed in silico analysis of chromatin assembly factor-1 (CAF-1) family in human malaria parasite Plasmodium falciparum. Our analysis revealed five chromatin assembly factor-1 genes in P. falciparum (PfCAF-1) and the PfCAF-1 family was divided into two classes where, Class A belongs to the CAF-1 complex and others are kept in Class B. For comparative studies, orthologs of PfCAF-1 family were identified across 53 eukaryotic species and evolutionary relationships were drawn for different CAF-1 subfamilies. The phylogenetic analysis revealed grouping of evolutionary-related species together, although, divergence was observed in branching pattern. A detailed analysis of domain composition highlighted species-specific features viz. species-specific KDDS repeats of 84 amino acids were identified in PfCAF-1A whereas, members of CAF-1C/RbAp48 and RbAp46 subfamily exhibited least variation in size and domain composition. The qRT-PCR analysis revealed upregulation of PfCAF-1 members in trophozoite or schizont stage. Furthermore, a comparative expression analysis of the available transcriptome and proteome data along with qRT-PCR analysis revealed mixed expression patterns (coordination as well as non-coordination between different studies). Protein-protein interaction network analyses of PfCAF-1 family were carried out highlighting important complexes based on interologs. The PfRbAp48 was found to be highly connected with a total of 108 PPIs followed by PfRbAp46. The results unravel insights into the PfCAF-1 family and identify unique features, thus opening new perspectives for further targeted developments to understand and combat malaria menace.