Centrioles are microtubule-based cellular structures present in most human cells that build centrosomes and cilia. Proliferating cells have only two centrosomes and this number is stringently maintained through the temporally and spatially controlled processes of centriole assembly and segregation. The assembly of new centrioles begins in early S phase and ends in the third G1 phase from their initiation. This lengthy process of centriole assembly from their initiation to their maturation is characterized by numerous structural and still poorly understood biochemical changes, which occur in synchrony with the progression of cells through three consecutive cell cycles. As a result, proliferating cells contain three structurally, biochemically, and functionally distinct types of centrioles: procentrioles, daughter centrioles, and mother centrioles. This age difference is critical for proper centrosome and cilia function. Here we discuss the centriole assembly process as it occurs in somatic cycling human cells with a focus on the structural, biochemical, and functional characteristics of centrioles of different ages.

https://www.mdpi.com/2073-4409/9/6/1429/pdf

With Age Comes Maturity: Biochemical and Structural Transformation of a Human Centriole in the Making

Primary microcephaly (MCPH) is a neurodevelopmental disorder characterized by small brain size with mental retardation. CPAP/CENPJ, a known microcephaly gene, plays a key role in centriole biogenesis. Here, we generated a previously unreported conditional knockout allele in the mouse Cpap gene. Our results showed that conditional Cpap deletion preferentially induces formation of monopolar spindles in radial glia progenitors (RGPs) and causes robust apoptosis that severely disrupts embryonic brains. Interestingly, microcephalic brains with reduced apoptosis are detected in the conditional Cpap gene-deleted mice, which lose only one allele of p53, while simultaneous removal of p53 and Cpap rescues RGPs death. Furthermore, Cpap deletion leads to cilia loss, RGPs mislocalization, junctional integrity disruption, massive heterotopia, and severe cerebellar hypoplasia. Together, these findings indicate that complete CPAP loss leads to severe and complex phenotypes in developing mouse brain, and provide new insights into the causes of MCPH.

/pdf/loss-of-cpap-in-developing-mouse-brain-and-its-functional-5d83vzp7oi.pdf

Loss of CPAP in developing mouse brain and its functional implication in human primary microcephaly.

Cells inherit two centrioles, the older of which is uniquely capable of generating a cilium. Using proteomics and superresolved imaging, we identify a module that we term DISCO (distal centriole complex). The DISCO components CEP90, MNR, and OFD1 underlie human ciliopathies. This complex localizes to both distal centrioles and centriolar satellites, proteinaceous granules surrounding centrioles. Cells and mice lacking CEP90 or MNR do not generate cilia, fail to assemble distal appendages, and do not transduce Hedgehog signals. Disrupting the satellite pools does not affect distal appendage assembly, indicating that it is the centriolar populations of MNR and CEP90 that are critical for ciliogenesis. CEP90 recruits the most proximal known distal appendage component, CEP83, to root distal appendage formation, an early step in ciliogenesis. In addition, MNR, but not CEP90, restricts centriolar length by recruiting OFD1. We conclude that DISCO acts at the distal centriole to support ciliogenesis by restraining centriole length and assembling distal appendages, defects in which cause human ciliopathies.

A ciliopathy complex builds distal appendages to initiate ciliogenesis.

Disease-Associated Mutations in CEP120 Destabilize the Protein and Impair Ciliogenesis.

Centrosomes are the main microtubule-organizing centres, playing essential roles in the organization of the cytoskeleton during interphase, and in the mitotic spindle, which controls chromosome segregation, during cell division. Centrosomes also act as the basal body of cilia, regulating cilium length and affecting extracellular signal reception as well as the integration of intracellular signalling pathways. Centrosomes are self-replicative and duplicate once every cell cycle to generate two centrosomes. The core support structure of the centrosome consists of two molecularly distinct centrioles. The mother (mature) centriole exhibits accessory appendages and is surrounded by both pericentriolar material and centriolar satellites, structures that the daughter (immature) centriole lacks. In this Review, we discuss what is currently known about centrosome duplication, its dialogue with the cell cycle and the sequential acquisition of specific components during centriole maturation. We also describe our current understanding of the mature centriolar structures that are required to build a cilium. Altogether, the built-in centrosome asymmetries that stem from the two centrosomes inheriting molecularly different centrioles sets the foundation for cell division being an intrinsically asymmetric process.

/pdf/centrosome-maturation-in-tune-with-the-cell-cycle-2zzbddh1.pdf

Centrosome maturation - in tune with the cell cycle.

Centrosomal protein 120 (CEP120) was originally identified as a daughter centriole-enriched protein that participates in centriole elongation. Recent studies showed that CEP120 gene mutations cause complex ciliopathy phenotypes in humans, including Joubert syndrome and Jeune asphyxiating thoracic dystrophy, suggesting that CEP120 plays an additional role in ciliogenesis. To investigate the potential roles of CEP120 in centriole elongation and cilia formation, we knocked out the CEP120 gene in p53-deficient RPE1 cells using the CRISPR/Cas9 editing system, and performed various analyses. We herein report that loss of CEP120 produces short centrioles with no apparent distal and subdistal appendages. CEP120 knockout was also associated with defective centriole elongation, impaired recruitment of C2CD3 and Talpid3 to the distal ends of centrioles, and consequent defects in centriole appendage assembly and cilia formation. Interestingly, wild-type CEP120 interacts with C2CD3 and Talpid3, whereas a disease-associated CEP120 mutant (I975S) has a low affinity for C2CD3 binding and perturbs cilia assembly. Together, our findings reveal a novel role of CEP120 in ciliogenesis by showing that it interacts with C2CD3 and Talpid3 to assemble centriole appendages and by illuminating the molecular mechanism through which the CEP120 (I975S) mutation causes complex ciliopathies.

/pdf/cep120-interacts-with-c2cd3-and-talpid3-and-is-required-for-ey5fuled32.pdf

CEP120 interacts with C2CD3 and Talpid3 and is required for centriole appendage assembly and ciliogenesis

The transcription factor Ron1 is required for chitin metabolism, asexual development and pathogenicity in Beauveria bassiana, an entomopathogenic fungus.

In this study, we report the roles of CDA family in entomopathogenic fungus B. bassiana. Our results indicated that CDA modulates asexual development and regulates fungal virulence by altering chitin deacetylation and metabolic capacity. CDA affected the biological control potential and life history of B. bassiana by affecting its parasitic and saprophytic life. These findings provide novel insights into the roles of multiple CDA paralogues existing in fungal biocontrol agents. ABSTRACT As an important chitin-modifying enzyme, chitin deacetylase (CDA) has been characterized in many fungi, but its function in the entomopathogenic fungus Beauveria bassiana remains unclear. Three CDAs with conserved domains of the carbohydrate esterase 4 (CE-4) family were identified in B. bassiana. Disruption of CDA1 resulted in growth restriction of the fungus on medium with chitin as a carbon source or without a carbon source. Deletion of CDA1 and CDA2 led to defects in fungal conidial formation and conidial vitality compared with those of the wild type (WT), and the conidial yield decreased by 25.81% to 47.68%. Inactivation of three CDA genes resulted in a decrease of 20.23% to 27% in the blastospore yield. ΔCDA1 and ΔCDA3 showed 29.33% and 23.34% reductions in cuticular infection virulence, respectively. However, the CDA family may not contribute to hemocoel infection virulence. Additionally, the sporulation of the insect carcass showed that the three gene deletion mutants were 68.45%, 63.84%, and 56.65% less than WT. Penetration experiments with cicada wings and enzyme activity assays were used to further explore the effect of the fungus on chitin metabolism after gene deletion. Although the three gene deletion mutants penetrated the cicada wings successfully and continued to grow on the underlying medium, their colony sizes were reduced by 29.12% to 47.76%. The CDA enzyme activity of ΔCDA1 and ΔCDA3 decreased by 84.76% and 83.04%, respectively. These data showed that members of the CDA family play a different role in fungal growth, conidial quality, and virulence. IMPORTANCE In this study, we report the roles of CDA family in entomopathogenic fungus B. bassiana. Our results indicated that CDA modulates asexual development and regulates fungal virulence by altering chitin deacetylation and metabolic capacity. CDA affected the biological control potential and life history of B. bassiana by affecting its parasitic and saprophytic life. These findings provide novel insights into the roles of multiple CDA paralogues existing in fungal biocontrol agents.

https://doi.org/10.1128/spectrum.04748-22

Three Chitin Deacetylase Family Members of Beauveria bassiana Modulate Asexual Reproduction and Virulence of Fungi by Mediating Chitin Metabolism and Affect Fungal Parasitism and Saprophytic Life

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

/pdf/author-correction-cep120-interacts-with-c2cd3-and-talpid3-4xs7xubrxd.pdf

Author Correction: CEP120 interacts with C2CD3 and Talpid3 and is required for centriole appendage assembly and ciliogenesis

The histone acetyltransferase Gcn5 ortholog contributes significantly to the growth and development of various fungi. In this study, we found that Ada2 and Ada3 have critical regulatory effects on Gcn5 enzyme activity and influence the acetylation of histone H3. ABSTRACT The histone acetyltransferase (HAT) Gcn5 ortholog is essential for a variety of fungi. Here, we characterize the roles of Ada2 and Ada3, which are functionally linked to Gcn5, in the insect-pathogenic fungus Beauveria bassiana. Loss of Ada2 and Ada3 led to severe hyphal growth defects on rich and minimal media and drastic decreases in blastospore yield and conidiation capacity, with abnormal conidia-producing structures. ΔAda2 and ΔAda3 exhibited a delay in conidial germination and increased sensitivity to multiple chemical stresses and heat shock. Nearly all their pathogenicity was lost, and their ability to secrete extracellular enzymes, Pr1 proteases and chitinases for cuticle degradation was reduced. A yeast two-hybrid assay demonstrated that Ada2 binds to Ada3 and directly interacts with Gcn5, confirming the existence of a yeast-like Ada3-Ada2-Gcn5 HAT complex in this fungus. Additionally, deletion of the Ada genes reduced the activity of Gcn5, especially in the ΔAda2 strain, which was consistent with the acetylation level of histone H3 determined by Western blotting. These results illustrate the dependence of Gcn5 enzyme activity on Ada2 and Ada3 in fungal hyphal growth, asexual development, multiple stress responses, and pathogenicity in B. bassiana. IMPORTANCE The histone acetyltransferase Gcn5 ortholog contributes significantly to the growth and development of various fungi. In this study, we found that Ada2 and Ada3 have critical regulatory effects on Gcn5 enzyme activity and influence the acetylation of histone H3. Deletion of Ada2 or Ada3 decreased the fungal growth rate and asexual conidial yield and increased susceptibility to multiple stresses in Beauveria bassiana. Importantly, Ada genes are vital virulence factors, and their deletion caused the most virulence loss, mainly by inhibiting the activity of a series of hydrolytic enzymes and the dimorphic transition ability. These findings provide a new perspective on the function of the Gcn5 acetyltransferase complex in pathogens.

https://doi.org/10.1128/spectrum.00281-23

Jia-Hua Liu

Papers

CEP120 interacts with C2CD3 and Talpid3 and is required for centriole appendage assembly and ciliogenesis

The transcription factor Ron1 is required for chitin metabolism, asexual development and pathogenicity in Beauveria bassiana, an entomopathogenic fungus.

Three Chitin Deacetylase Family Members of Beauveria bassiana Modulate Asexual Reproduction and Virulence of Fungi by Mediating Chitin Metabolism and Affect Fungal Parasitism and Saprophytic Life

Author Correction: CEP120 interacts with C2CD3 and Talpid3 and is required for centriole appendage assembly and ciliogenesis

Ada2 and Ada3 Regulate Hyphal Growth, Asexual Development, and Pathogenicity in Beauveria bassiana by Maintaining Gcn5 Acetyltransferase Activity