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Centrioles are polarized microtubule-based structures with appendages at their distal end that are essential for cilia formation and function. The protein C2CD3 is critical for distal appendage assembly, with mutations linked to orofaciodigital syndrome and other ciliopathies. However, its precise molecular role in appendage recruitment remains unclear. Using ultrastructure expansion microscopy (U-ExM) and iterative U-ExM on human cells, together with in situ cryo-electron tomography (cryo-ET) on mouse tissues, we reveal that C2CD3 adopts a radially symmetric 9-fold organization within the centriole’s distal lumen. We show that the C-terminal region of C2CD3 localizes close to a ~100 nm luminal ring structure consisting of ~27 nodes, while its N-terminal region localizes close to a hook-like structure that attaches to the A-microtubule as it extends from the centriole interior to exterior. This hook structure is adjacent to the DISCO complex (MNR/CEP90/OFD1), which marks future appendage sites. C2CD3 depletion disrupts not only the recruitment of the DISCO complex via direct interaction with MNR but also destabilizes the luminal ring network composed of C2CD3/SFI1/centrin-2/CEP135/NA14, as well as the distal microtubule tip protein CEP162. This reveals an intricate “in-to-out” molecular hub connecting the centriolar lumen, distal microtubule cap, and appendages. Although C2CD3 loss results in shorter centrioles and appendage defects, key structural elements remain intact, permitting continued centriole duplication. We propose that C2CD3 forms the luminal ring structure and extends radially to the space between triplet microtubules, functioning as an architectural hub that scaffolds the distal end of the centriole, orchestrating its assembly and directing appendage formation.

Centrioles are polarized microtubule-based structures with appendages at their distal end that are essential for cilia formation and function. The protein C2CD3 is critical for distal appendage assembly, with mutations linked to orofaciodigital syndrome and other ciliopathies. However, its precise molecular role in appendage recruitment remains unclear. Using Ultrastructure Expansion Microscopy (U-ExM), iterative U-ExM, and cryo-electron tomography (cryo-ET), we reveal that C2CD3 adopts a radially symmetric 9-fold organization within the centriole's distal lumen. We show that the C-terminal region of C2CD3 localizes close to a ~100 nm luminal ring structure consisting of ~27 nodes, while its N-terminal region localizes close to a hook-like structure that attaches to the A-microtubule as it extends from the centriole interior to exterior. This hook structure is adjacent to the DISCO complex (MNR/CEP90/OFD1), which marks future appendage sites. C2CD3 depletion disrupts not only the recruitment of the DISCO complex via direct interaction with MNR but also destabilizes the luminal ring network composed of C2CD3/SFI1/centrin-2/CEP135/NA14, as well as the distal microtubule tip protein CEP162. This reveals an intricate \"in-to-out\" molecular hub connecting the centriolar lumen, distal microtubule cap, and appendages. Although C2CD3 loss results in shorter centrioles and appendage defects, key structural elements remain intact, permitting continued centriole duplication. We propose that C2CD3 forms the luminal ring structure and extends radially to the space between triplet microtubules, functioning as an architectural hub that scaffolds the distal end of the centriole, orchestrating its assembly and directing appendage formation.

Malate (2-hydroxysuccinic acid) and tartrate (2,3-dihydroxysuccinic acid) are chiral substrates; the former existing in two enantiomeric forms (R and S) while the latter exists as three stereoisomers (R,R; S,S; and R,S). Dehydration by stereospecific hydrogen abstraction and anti- elimination of the hydroxyl group yield the achiral products fumarate and oxaloacetate, respectively. Class-I fumarate hydratase (FH) and L-tartrate dehydratase (L-TTD) are two highly conserved enzymes belonging to the iron-sulfur cluster hydrolyase family of enzymes that catalyze reactions on specific stereoisomers of malate and tartrate. FH from Methanocaldococcus jannaschii accepts only S-malate and S,S-tartrate as substrates while the structurally similar L-TTD from Escherichia coli accepts only R-malate and R,R-tartrate as substrates. Phylogenetic analysis reveals a common evolutionary origin of L-TTDs and two-subunit archaeal FHs suggesting a divergence during evolution that may have led to the switch in substrate stereospecificity preference. Due to the high conservation of their sequences, a molecular basis for switch in stereospecificity is not evident from analysis of crystal structures of FH and predicted structure of L-TTD. The switch in enantiomer preference may be rationalised by invoking conformational plasticity of the amino acids interacting with the substrate, together with substrate reorientation and conformer selection about the C2-C3 bond of the dicarboxylic acid substrates. Although classical models of enzyme-substrate binding are insufficient to explain such a phenomenon, the enantiomer superposition model suggests that a minor reorientation in the active site residues could lead to the switch in substrate stereospecificity.

Succinimide (SNN), an intermediate formed during asparaginyl deamidation or aspartyl dehydration in proteins, is generally hydrolysis-prone, leading to isomerization to L/D α/β-aspartyl residue, with the latter being considered deleterious to protein structure and function. An unusually stable SNN-mediated conformational rigidity through restriction of the backbone dihedral angle, ψ, enhances the thermostability of glutamine amidotransferase (GATase) from Methanocaldococcus jannaschii (Mj). Although several structural features involved in maintaining a stable SNN and imparting SNN-mediated thermostability have been identified in MjGATase, the residues in the protein that catalyse the rapid and complete conversion of Asn109 to SNN remain unknown. Here, we investigated several site-directed mutants of MjGATase for their ability to retain Asn109 side chain in the unmodified form. Mass spectrometric analysis of 10 single mutants enabled the identification of residues that impacted the proportion of SNN and Asn population in the protein sample. This led to the generation of two double mutants that retained intact Asn109 side chain as observed in the mass spectra and crystal structures. These mutants with intact Asn residue at position 109, displayed lower thermal stability than the protein with the SNN modification. Further understanding of the deprotonation mechanism was addressed using QM/MM MD metadynamics simulations. Stable succinimide (SNN) arising from deamidation of Asn109 residue imparts hyperthermostability to MjGATase. Examination of the structure of MjGATase suggests neighbouring residues playing possible roles in deamidation and cyclization. Examination by LC-MS of single site-directed mutants of residues contacting SNN revealed varied levels of intact Asn109 enabling generation of double mutants with complete absence of deamidation. Presence of intact Asn109 confirmed by X-ray crystallography highlights the role of Y158, D110, and K151 in mediating SNN formation. QM/MM MD metadynamics simulations support experimental findings.

PurposeThe COVID-19 pandemic has resulted in countries reacting differently to an ongoing crisis. Latent to this reaction mechanism is the inherent cultural characteristics of each society resulting in differential responses to the epidemic spread. In this study, the moderated moderation role of culture, on information dissemination by media during epidemic recovery-phase has been investigated.Design/methodology/approachHofstede’s cultural factors are hypothesized to moderate the moderating effect of free-liberal media on the relationship of COVID-19 recovery rate and human mobility. Panel regression model, using mobility data and recovery rate across 95 countries for a period of 170 days has been preferred to test the hypotheses. The results are further substantiated using factor wise interaction plots and slope difference analysis.FindingsThe findings suggest that societies with high power distance and masculinity scores strengthen the impact of media on the relationship between COVID-19 recovery rate and mobility whereas, high individualistic and long-term orientation societies weaken the same effect. However, similar conclusions were not confirmed for uncertainty avoidance. Cross-cultural impact, as elucidated by this study, forms a crucial element in policy formulation on epidemic control by indigenous Governing bodies.Originality/valueWhile most of the studies emphasizing on cultural characteristics of a society in an epidemic situation covers the growth phase of infection, This research talks about the recovery-phase of the epidemic and the effect of culture.Peer reviewThe peer review history for this article is available at: https://publons.com/publon/10.1108/IJSE-04-2023-0314

Background Charcot-Marie-Tooth (CMT) is a clinically, electro-physiologically, and genetically heterogenous group of muscle disease which is also known as hereditary motor and sensory neuropathy. Autosomal recessive forms of CMT type 4A have been reported with either homozygous or compound heterozygous mutations in a gene that encodes ganglioside-induced differentiation-associated protein-1 (GDAP1). GDAP1 is located on 8q21, and plays a major role in ganglioside differentiation and Schwann cell function, as well as regulates neuronal and axonal development. Case presentation In this study, we recruited a consanguineous south Indian family with an affected patient, an unaffected sibling, and the mother. The patient was affected with progressive weakness in the lower and upper limbs, atrophy of small muscles of the foot and hands, club shaped hands, steppage gait, hoarseness, and decreased muscle tone. His nerve biopsy examination revealed peripheral nerve demyelination and nerve conduction testing confirmed a reduction in nerve activities, while MRI showed mild degenerative changes in the cervical spine. Further, targeted exome sequencing (TES) and copy number variation analysis were performed on the patient. TES identified a compound heterozygous mutation that includes a missense mutation and a 3'UTR mutation (NM_018972.4: c.413A > G:p.His138Arg; g.74488790C > A:c.*29C > A, respectively) in GDAP1. The missense change is not reported in available public databases, while the UTR variant is seen only in the South Asian population in gnomAD (allele frequency = 0.00002). Multiple in silico prediction tools show that the missense mutation is damaging. Subsequently, in silico protein modeling, phylogenetic conservation analysis, and the impact of the mutation on the canonical transcript have also been performed. The compound heterozygous mutation was confirmed in the patient by PCR-Sanger sequencing and was shown to segregate within the family. Conclusions The combined results support the fact that these two mutations in GDAP1 link the genotype-phenotype correlation in the family. This will help the family in genetic testing, counseling, and early diagnosis. Our findings support expanded phenotypic characterization along with the genetic spectrum of GDAP1 mutations in CMT type4A in the Indian population.

A 60-year-old gentleman presented with a large swelling on the left side of his neck which was diagnosed as a metastatic cervical lymph node. It infiltrated the platysma muscle and caused partial contracture. This is known as Platysma sign. Presence of this sign indicates extracapsular spread and shows poor prognosis.

Charcot-Marie-Tooth (CMT) is a clinically, electro-physiologically, and genetically heterogenous group of muscle disease which is also known as hereditary motor and sensory neuropathy. Autosomal recessive forms of CMT type 4A have been reported with either homozygous or compound heterozygous mutations in a gene that encodes ganglioside-induced differentiation-associated protein-1 (GDAP1). GDAP1 is located on 8q21, and plays a major role in ganglioside differentiation and Schwann cell function, as well as regulates neuronal and axonal development. The combined results support the fact that these two mutations in GDAP1 link the genotype-phenotype correlation in the family. This will help the family in genetic testing, counseling, and early diagnosis. Our findings support expanded phenotypic characterization along with the genetic spectrum of GDAP1 mutations in CMT type4A in the Indian population.