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European Journal of Human Genetics :... Aug 2023High-throughput sequencing has become a standard first-tier approach for both diagnostics and research-based genetic testing. Consequently, this hypothesis-free... (Review)
Review
High-throughput sequencing has become a standard first-tier approach for both diagnostics and research-based genetic testing. Consequently, this hypothesis-free testing manner has revealed the true breadth of clinical features for many established genetic disorders, including Meier-Gorlin syndrome (MGORS). Previously known as ear-patella short stature syndrome, MGORS is characterized by growth delay, microtia, and patella hypo/aplasia, as well as genital abnormalities, and breast agenesis in females. Following the initial identification of genetic causes in 2011, a total of 13 genes have been identified to date associated with MGORS. In this review, we summarise the genetic and clinical findings of each gene associated with MGORS and highlight molecular insights that have been made through studying patient variants. We note interesting observations arising across this group of genes as the number of patients has increased, such as the unusually high number of synonymous variants affecting splicing in CDC45 and a subgroup of genes that also cause craniosynostosis. We focus on the complicated molecular genetics for DONSON, where we examine potential genotype-phenotype patterns using the first 3D structural model of DONSON. The canonical role of all proteins associated with MGORS are involved in different stages of DNA replication and in addition to summarising how patient variants impact on this process, we discuss the potential contribution of non-canonical roles of these proteins to the pathophysiology of MGORS.
Topics: Female; Humans; Congenital Microtia; Patella; Growth Disorders; Micrognathism
PubMed: 37059840
DOI: 10.1038/s41431-023-01359-z -
MBio Aug 2023The cellular processes that support human coronavirus replication and contribute to the pathogenesis of severe disease remain incompletely understood. Many viruses,...
The cellular processes that support human coronavirus replication and contribute to the pathogenesis of severe disease remain incompletely understood. Many viruses, including coronaviruses, cause endoplasmic reticulum (ER) stress during infection. IRE1α is a component of the cellular response to ER stress that initiates non-conventional splicing of mRNA. Spliced encodes a transcription factor that induces the expression of ER-related targets. Activation of the IRE1α-XBP1 pathway occurs in association with risk factors for severe human coronavirus infection. In this study, we found that the human coronaviruses HCoV-OC43 (human coronavirus OC43) and SARS-CoV-2 (severe acute respiratory syndrome coronavirus-2) both robustly activate the IRE1α-XBP1 branch of the unfolded protein response in cultured cells. Using IRE1α nuclease inhibitors and genetic knockdown of IRE1α and XBP1, we found that these host factors are required for optimal replication of both viruses. Our data suggest that IRE1α supports infection downstream of initial viral attachment and entry. In addition, we found that ER stress-inducing conditions are sufficient to enhance human coronavirus replication. Furthermore, we found markedly increased XBP1 in circulation in human patients with severe coronavirus disease 2019 (COVID-19). Together, these results demonstrate the importance of IRE1α and XBP1 for human coronavirus infection. IMPORTANCE There is a critical need to understand the cellular processes co-opted during human coronavirus replication, with an emphasis on identifying mechanisms underlying severe disease and potential therapeutic targets. Here, we demonstrate that the host proteins IRE1α and XBP1 are required for robust infection by the human coronaviruses, SARS-CoV-2 and HCoV-OC43. IRE1α and XBP1 participate in the cellular response to ER stress and are activated during conditions that predispose to severe COVID-19. We found enhanced viral replication with exogenous IRE1α activation, and evidence that this pathway is activated in humans during severe COVID-19. Together, these results demonstrate the importance of IRE1α and XBP1 for human coronavirus infection.
Topics: Humans; Protein Serine-Threonine Kinases; Endoribonucleases; COVID-19; SARS-CoV-2; Unfolded Protein Response; Endoplasmic Reticulum Stress; X-Box Binding Protein 1
PubMed: 37306512
DOI: 10.1128/mbio.00540-23 -
Nano Convergence Sep 2023Nanozymes mimic the function of enzymes, which drive essential intracellular chemical reactions that govern biological processes. They efficiently generate or degrade... (Review)
Review
Nanozymes mimic the function of enzymes, which drive essential intracellular chemical reactions that govern biological processes. They efficiently generate or degrade specific biomolecules that can initiate or inhibit biological processes, regulating cellular behaviors. Two approaches for utilizing nanozymes in intracellular chemistry have been reported. Biomimetic catalysis replicates the identical reactions of natural enzymes, and bioorthogonal catalysis enables chemistries inaccessible in cells. Various nanozymes based on nanomaterials and catalytic metals are employed to attain intended specific catalysis in cells either to mimic the enzymatic mechanism and kinetics or expand inaccessible chemistries. Each nanozyme approach has its own intrinsic advantages and limitations, making them complementary for diverse and specific applications. This review summarizes the strategies for intracellular catalysis and applications of biomimetic and bioorthogonal nanozymes, including a discussion of their limitations and future research directions.
PubMed: 37695365
DOI: 10.1186/s40580-023-00390-6 -
The Journal of Infectious Diseases Nov 2023Ebola virus (EBOV) disease is marked by rapid virus replication and spread. EBOV enters the cell by macropinocytosis and replicates in the cytoplasm, and nascent virions...
Ebola virus (EBOV) disease is marked by rapid virus replication and spread. EBOV enters the cell by macropinocytosis and replicates in the cytoplasm, and nascent virions egress from the cell surface to infect neighboring cells. Here, we show that EBOV uses an alternate route to disseminate: tunneling nanotubes (TNTs). TNTs, an actin-based long-range intercellular communication system, allows for direct exchange of cytosolic constituents between cells. Using live, scanning electron, and high-resolution quantitative 3-dimensional microscopy, we show that EBOV infection of primary human cells results in the enhanced formation of TNTs containing viral nucleocapsids. TNTs promote the intercellular transfer of nucleocapsids in the absence of live virus, and virus could replicate in cells devoid of entry factors after initial stall. Our studies suggest an alternate model of EBOV dissemination within the host, laying the groundwork for further investigations into the pathogenesis of filoviruses and, importantly, stimulating new areas of antiviral design.
Topics: Humans; Ebolavirus; Hemorrhagic Fever, Ebola; Nanotubes; Cell Communication
PubMed: 37723997
DOI: 10.1093/infdis/jiad400 -
EcoSal Plus Dec 2023To preserve the integrity of their genome, bacteria rely on several genome maintenance mechanisms that are co-ordinated with the cell cycle. All members of the family... (Review)
Review
To preserve the integrity of their genome, bacteria rely on several genome maintenance mechanisms that are co-ordinated with the cell cycle. All members of the family have a bipartite genome consisting of a primary chromosome (Chr1) homologous to the single chromosome of other bacteria such as and a secondary chromosome (Chr2) acquired by a common ancestor as a plasmid. In this review, we present our current understanding of genome maintenance in , which is the best-studied model for bacteria with multi-partite genomes. After a brief overview on the diversity of genomic architecture, we describe the specific, common, and co-ordinated mechanisms that control the replication and segregation of the two chromosomes of . Particular attention is given to the unique checkpoint mechanism that synchronizes Chr1 and Chr2 replication.
Topics: DNA Replication; Bacterial Proteins; Chromosomes, Bacterial; Cell Cycle; Vibrio cholerae; Bacteria
PubMed: 38277776
DOI: 10.1128/ecosalplus.esp-0008-2022 -
The Journal of Allergy and Clinical... Jul 2023Severe congenital neutropenia presents with recurrent infections early in life as a result of arrested granulopoiesis. Multiple genetic defects are known to block...
BACKGROUND
Severe congenital neutropenia presents with recurrent infections early in life as a result of arrested granulopoiesis. Multiple genetic defects are known to block granulocyte differentiation; however, a genetic cause remains unknown in approximately 40% of cases.
OBJECTIVE
We aimed to characterize a patient with severe congenital neutropenia and syndromic features without a genetic diagnosis.
METHODS
Whole exome sequencing results were validated using flow cytometry, Western blotting, coimmunoprecipitation, quantitative PCR, cell cycle and proliferation analysis of lymphocytes and fibroblasts and granulocytic differentiation of primary CD34 and HL-60 cells.
RESULTS
We identified a homozygous missense mutation in DBF4 in a patient with mild extra-uterine growth retardation, facial dysmorphism and severe congenital neutropenia. DBF4 is the regulatory subunit of the CDC7 kinase, together known as DBF4-dependent kinase (DDK), the complex essential for DNA replication initiation. The DBF4 variant demonstrated impaired ability to bind CDC7, resulting in decreased DDK-mediated phosphorylation, defective S-phase entry and progression and impaired differentiation of granulocytes associated with activation of the p53-p21 pathway. The introduction of wild-type DBF4 into patient CD34 cells rescued the promyelocyte differentiation arrest.
CONCLUSION
Hypomorphic DBF4 mutation causes autosomal-recessive severe congenital neutropenia with syndromic features.
Topics: Humans; Cell Cycle Proteins; Protein Serine-Threonine Kinases; Saccharomyces cerevisiae Proteins; Mutation; Phosphorylation
PubMed: 36841265
DOI: 10.1016/j.jaci.2023.02.016 -
Life (Basel, Switzerland) Feb 2024To track down the possible roots of life, various models for the initial living system composed of different combinations of the three extant biopolymers, RNA, DNA, and... (Review)
Review
To track down the possible roots of life, various models for the initial living system composed of different combinations of the three extant biopolymers, RNA, DNA, and proteins, are presented. The suitability of each molecular set is assessed according to its ability to emerge autonomously, sustain, and evolve continuously towards life as we know it. The analysis incorporates current biological knowledge gained from high-resolution structural data and large sequence datasets, together with experimental results concerned with RNA replication and with the activity demonstrated by standalone constructs of the ribosomal Peptidyl Transferase Center region. The scrutiny excludes the DNA-protein combination and assigns negligible likelihood to the existence of an RNA-DNA world, as well as to an RNA world that contained a replicase made of RNA. It points to the precedence of an RNA-protein system, whose model of emergence suggests specific processes whereby a coded proto-ribosome ribozyme, specifically aminoacylated proto-tRNAs and a proto-polymerase enzyme, could have autonomously emerged, cross-catalyzing the formation of each other. This molecular set constitutes a feasible starting point for a continuous evolutionary path, proceeding via natural processes from the inanimate matter towards life as we know it.
PubMed: 38398786
DOI: 10.3390/life14020277 -
Frontiers in Microbiology 2023Although the mechanism of DNA replication initiation has been investigated for over 50 years, many important discoveries have been made related to this process in... (Review)
Review
Although the mechanism of DNA replication initiation has been investigated for over 50 years, many important discoveries have been made related to this process in recent years. In this mini-review, we discuss the current state of knowledge concerning the structure of the origin region in bacterial chromosomes and plasmids, recently discovered motifs recognized by replication initiator proteins, and proposed in the literature models describing initial origin opening. We review structures of nucleoprotein complexes formed by replication initiators at chromosomal and plasmid replication origins and discuss their functional implications. We also discuss future research challenges in this field.
PubMed: 38249469
DOI: 10.3389/fmicb.2023.1328842 -
Cells Jul 2023The mammalian Ccr4-Not complex, carbon catabolite repression 4 (Ccr4)-negative on TATA-less (Not), is a large, highly conserved, multifunctional assembly of proteins...
The mammalian Ccr4-Not complex, carbon catabolite repression 4 (Ccr4)-negative on TATA-less (Not), is a large, highly conserved, multifunctional assembly of proteins that acts at different cellular levels to regulate gene expression. It is involved in the control of the cell cycle, chromatin modification, activation and inhibition of transcription initiation, control of transcription elongation, RNA export, and nuclear RNA surveillance; the Ccr4-Not complex also plays a central role in the regulation of mRNA decay. Growing evidence suggests that gene transcription has a vital role in shaping the landscape of genome replication and is also a potent source of replication stress and genome instability. Here, we have examined the effects of the inactivation of the Ccr4-Not complex, via the depletion of the scaffold subunit CNOT1, on DNA replication and genome integrity in mammalian cells. In CNOT1-depleted cells, the elevated expression of the general transcription factor TATA-box binding protein (TBP) leads to increased RNA synthesis, which, together with R-loop accumulation, results in replication fork slowing, DNA damage, and senescence. Furthermore, we have shown that the stability of TBP mRNA increases in the absence of CNOT1, which may explain its elevated protein expression in CNOT1-depleted cells. Finally, we have shown the activation of mitogen-activated protein kinase signalling as evidenced by ERK1/2 phosphorylation in the absence of CNOT1, which may be responsible for the observed cell cycle arrest at the border of G1/S.
Topics: Animals; Transcription Factors; Catabolite Repression; RNA; RNA, Messenger; Genomic Instability; Mammals
PubMed: 37508532
DOI: 10.3390/cells12141868 -
The Journal of Biological Chemistry Nov 20233D chromatin organization plays a critical role in regulating gene expression, DNA replication, recombination, and repair. While initially discovered for its role in...
3D chromatin organization plays a critical role in regulating gene expression, DNA replication, recombination, and repair. While initially discovered for its role in sister chromatid cohesion, emerging evidence suggests that the cohesin complex (SMC1, SMC3, RAD21, and SA1/SA2), facilitated by NIPBL, mediates topologically associating domains and chromatin loops through DNA loop extrusion. However, information on how conformational changes of cohesin-NIPBL drive its loading onto DNA, initiation, and growth of DNA loops is still lacking. In this study, high-speed atomic force microscopy imaging reveals that cohesin-NIPBL captures DNA through arm extension, assisted by feet (shorter protrusions), and followed by transfer of DNA to its lower compartment (SMC heads, RAD21, SA1, and NIPBL). While binding at the lower compartment, arm extension leads to the capture of a second DNA segment and the initiation of a DNA loop that is independent of ATP hydrolysis. The feet are likely contributed by the C-terminal domains of SA1 and NIPBL and can transiently bind to DNA to facilitate the loading of the cohesin complex onto DNA. Furthermore, high-speed atomic force microscopy imaging reveals distinct forward and reverse DNA loop extrusion steps by cohesin-NIPBL. These results advance our understanding of cohesin by establishing direct experimental evidence for a multistep DNA-binding mechanism mediated by dynamic protein conformational changes.
Topics: Cell Cycle Proteins; Chromosomal Proteins, Non-Histone; DNA; Chromatin; Cohesins
PubMed: 37774974
DOI: 10.1016/j.jbc.2023.105296