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Background Congenital central hypoventilation syndrome (CCHS) is a rare autosomal dominant disease that is mainly caused by PHOX2B mutations. The purpose of this study is to analyze and summarize the clinical and genetic characteristics of CCHS patients in the Chinese population from our study and previous literature. Methods The potential pathogenic gene mutations of CCHS were identified and verified by next generation sequencing combined with Sanger sequencing, fluorescent probe PCR and capillary electrophoresis. The clinical characteristics and gene mutations of CCHS cases in Chinese population were summarized from our study and previous literature to explore the genotype–phenotype correlations. Results We identified 48 CCHS cases including three new cases from our report in China. Overall, 77.1% of the patients had PHOX2B polyalanine repeat expansion mutations (PARMs), and the remaining 22.9% had 10 distinct PHOX2B non‐polyalanine repeat expansion mutations (NPARMs). Compared to those with PARMs, patients with NPARMs were more likely to have premature birth (54.5% vs. 2.8%, p < 0.001) and lower birth weight (33.3% vs. 3.2%, p = 0.030), with statistical significance. The patients with PARMs were more likely to have cardiovascular defects (64.9% vs. 27.3%, p = 0.063), cerebral hemorrhage (29.7% vs. 9.1%, p = 0.322) and seizures (37.8% vs. 9.1%, p = 0.151) than those with NPARMs, with no statistical significance. Conclusions CCHS patients with PHOX2B NPARMs were more likely to have premature birth and low birth weight, while PHOX2B PARMs tended to be positively associated with the risk of cardiovascular defects, cerebral hemorrhage and seizures in Chinese population. The purpose of this study is to analyze the clinical data and mutations of three new cases, and to summarize the clinical and genetic characteristics of CCHS patients in the Chinese population from our study and previous literature. Our study not only expands the mutation spectrum of PHOX2B, but also improves our understanding of this disease CCHS.

The development of artificial spider silk with properties similar to native silk has been a challenging task in materials science. In this study, we use a microfluidic device to create continuous fibers based on recombinant MaSp2 spidroin. The strategy incorporates ion-induced liquid-liquid phase separation, pH-driven fibrillation, and shear-dependent induction of β-sheet formation. We find that a threshold shear stress of approximately 72 Pa is required for fiber formation, and that β-sheet formation is dependent on the presence of polyalanine blocks in the repetitive sequence. The MaSp2 fiber formed has a β-sheet content (29.2%) comparable to that of native dragline with a shear stress requirement of 111 Pa. Interestingly, the polyalanine blocks have limited influence on the occurrence of liquid-liquid phase separation and hierarchical structure. These results offer insights into the shear-induced crystallization and sequence-structure relationship of spider silk and have significant implications for the rational design of artificially spun fibers. Native spider silk has desirable mechanical properties, but these are challenging to replicate in an artificial material. Here, the authors report the use of a microfluidic system to create continuous fibers based on recombinant spidroin.

Growth of the eastern oyster Crassostrea virginica, a major aquaculture species in the USA, is highly variable and not well understood at molecular levels. As growth of mollusks is confined in shells constructed by the mantle, mantle transcriptomes of large (fast-growing) and small (slow-growing) eastern oysters were sequenced and compared in this study. Transcription was observed for 31,186 genes, among which 104 genes were differentially expressed between the large and small oysters, including 48 upregulated and 56 downregulated in large oysters. Differentially expressed genes (DEGs) included genes from diverse pathways highlighting the complexity of shell formation and growth regulations. Seventeen of the 48 upregulated DEGs were related to shell matrix formation, most of which were upregulated in large oysters, indicating that large oysters are more active in biomineralization and shell formation. Genomic and transcriptomic analyses identified 22 genes encoding novel polyalanine containing proteins (Pacps) with characteristic motifs for matrix function that are tandemly duplicated on one chromosome, all specifically expressed in mantle and at higher levels in large oysters, suggesting that these expanded Pacps play important roles in shell formation and growth. Analysis of sequence variation identified 244,964 SNPs with 328 associated with growth. This study provides novel candidate genes and markers for shell formation and growth, and suggests that genes related to shell formation are important for the complex regulation of growth in the eastern oyster and possibly other bivalve mollusks. Results of this study show that both transcriptional modulation and functional polymorphism are important in determining growth.

Background Synpolydactyly (SPD) is mainly caused by mutations of polyalanine expansion (PAE) in the transcription factor gene  HOXD13  and the involved cell types and signal pathway are still not clear possible pathways and single-cell expression characteristics of limb bud in  HOXD13 PAE mice was analyzed in this study. Method We investigated a previous study of a mouse model with SPD and conducted weighted gene co-expression network analysis (WGCNA) using a single-cell RNA sequencing dataset from limb bud cells of SPD mouse model of HOXD13  + 7A heterozygote. Results Analysis of WGCNA revealed that synpolydactyly-associated Hoxd13 PAEs alter the immune response and osteoclast differentiation, and enhance DNA replication. Bmp4 , Hand2 , Hoxd12 , Lnp , Prrx1 , Gmnn , and Cdc6 were found to play potentially key roles in synpolydactyly. Conclusions These findings evaluated the main genes related to SPD with PAE mutations in HOXD13 and advance our understanding of human limb development.

The mother, all her three affected sons as well as a niece and daughter of this woman’s sister (who was not tested), were all found to carry a mutation in their ARX gene RefSeq NM_139058.3: c.441_464dup p.Ala148_Ala155dup, resulting in a polyalanine repeat tract in the ARX protein. Figure 2 of Gras et al1 and figure 2 of Mattiske et al2 together appear to have shown that mutations in the ARX gene within and flanking the homeodomain as well as near the junction between exons 4-5 appear far more likely to be associated with severe/moderate phenotype in both females and males; both nonsense and missense mutations near the 5’ NH2 region, roughly before amino acid p.40 in exon one, appeared to be associated with normal phenotype in carrier females, but severe male phenotype; mutations near the 3’ COOH of the coding region, near and within the Aristaless domain also seem to be well tolerated by females but are typically associated with a severe/moderate phenotype in males; genotype-phenotype correlation appears to be more variable for mutations outside the above-mentioned regions. (II) From figure 2 of Gras et al (2024) showing positions of mutations from 5’ NH2 to 3’ COOH of the ARX gene and corresponding phenotypic severity in male and female patients.

The ribosome-associated quality-control (RQC) pathway degrades aberrant nascent polypeptides arising from ribosome stalling during translation. In mammals, the E3 ligase Pirh2 mediates the degradation of aberrant nascent polypeptides by targeting the C-terminal polyalanine degrons (polyAla/C-degrons). Here, we present the crystal structure of Pirh2 bound to the polyAla/C-degron, which shows that the N-terminal domain and the RING domain of Pirh2 form a narrow groove encapsulating the alanine residues of the polyAla/C-degron. Affinity measurements in vitro and global protein stability assays in cells further demonstrate that Pirh2 recognizes a C-terminal A/S-X-A-A motif for substrate degradation. Taken together, our study provides the molecular basis underlying polyAla/C-degron recognition by Pirh2 and expands the substrate recognition spectrum of Pirh2. Incompletely synthesized nascent polypeptides resulting from ribosome stalling during translation are under surveillance by ribosome-associated quality control. Here, the authors report the molecular mechanism by which the E3 ligase Pirh2 targets the polyalanine tail of aberrant nascent chains for degradation via the C-degron pathway.

Abnormal trinucleotide repeat expansions alter protein conformation causing malfunction and contribute to a significant number of incurable human diseases. Scarce structural insights available on disease-related homorepeat expansions hinder the design of effective therapeutics. Here, we present the dynamic structure of human PHOX2B C-terminal fragment, which contains the longest polyalanine segment known in mammals. The major α-helical conformation of the polyalanine tract is solely extended by polyalanine expansions in PHOX2B, which are responsible for most congenital central hypoventilation syndrome cases. However, polyalanine expansions in PHOX2B additionally promote nascent homorepeat conformations that trigger length-dependent phase transitions into solid condensates that capture wild-type PHOX2B. Remarkably, HSP70 and HSP90 chaperones specifically seize PHOX2B alternative conformations preventing phase transitions. The precise observation of emerging polymorphs in expanded PHOX2B postulates unbalanced phase transitions as distinct pathophysiological mechanisms in homorepeat expansion diseases, paving the way towards the search of therapeutics modulating biomolecular condensates in central hypoventilation syndrome. Here, the authors show that pathogenic mutations in the polyalanine expansions of PHOX2B promote nascent structural conformations that trigger irreversible phase transitions that arrest wild-type PHOX2B, disrupting function.

The general concept of tissue engineering is to restore biological function by replacing defective tissues with implantable, biocompatible, and easily handleable cell-laden scaffolds. In this study, osteoinductive and osteoconductive super paramagnetic Fe3O4 nanoparticles (MNP) and hydroxyapatite (HAP) nanoparticles were incorporated into a di-block copolymer based thermo-responsive hydrogel, methoxy(polyethylene glycol)-polyalanine (mPA), at various concentrations to afford composite, injectable hydrogels. Incorporating nanoparticles into the thermo-responsive hydrogel increased the complex viscosity and decreased the gelation temperature of the starting hydrogel. Functionally, the integration of inorganic nanoparticles modulated bio-markers of bone differentiation and enhanced bone mineralization. Moreover, this study adopted the emerging method of using either a supplementary static magnetic field (SMF) or a moving magnetic field to elicit biological response. These results demonstrate that combining external (magnet) and internal (scaffold) magnetisms is a promising approach for bone regeneration.

Polyalanine (polyA) disease-causative proteins with an expansion of alanine repeats can be aggregated. Although curative treatments for polyA diseases have not been explored, the dissociation of polyA aggregates likely reduces the cytotoxicity of polyA. Mid-infrared free electron laser (FEL) successfully dissociated multiple aggregates. However, whether the FEL dissociates polyA aggregates like other aggregates has not been tested. Here, we show that FEL at 6.1 μm experimentally weakened the extent of aggregation of a peptide with 13 alanine repeats (13A), and the irradiated 13A exerted lesser cytotoxicity to neuron-like cells than non-irradiated 13A. Then, we applied molecular dynamics (MD) simulation to follow the dissociation process by FEL. We successfully observed how the intermolecular β-sheet of polyA aggregates was dissociated and separated into monomers with helix structures upon FEL irradiation. After the dissociation by FEL, water molecules inhibited the reformation of polyA aggregates. We recently verified the same dissociation process using FEL-treated amyloid-β aggregates. Thus, a common mechanism underlies the dissociation of different protein aggregates that cause different diseases, polyA disease and Alzheimer’s disease. However, MD simulation indicated that polyA aggregates are less easily dissociated than amyloid-β aggregates and require longer laser irradiation due to hydrophobic alanine repeats.

Oculopharyngeal muscular dystrophy (OPMD) is a late-onset disorder characterized by progressive weakness and degeneration of specific muscles. OPMD is due to extension of a polyalanine tract in poly(A) binding protein nuclear 1 (PABPN1). Aggregation of the mutant protein in muscle nuclei is a hallmark of the disease. Previous transcriptomic analyses revealed the consistent deregulation of the ubiquitin-proteasome system (UPS) in OPMD animal models and patients, suggesting a role of this deregulation in OPMD pathogenesis. Subsequent studies proposed that UPS contribution to OPMD involved PABPN1 aggregation. Here, we use a Drosophila model of OPMD to address the functional importance of UPS deregulation in OPMD. Through genome-wide and targeted genetic screens we identify a large number of UPS components that are involved in OPMD. Half dosage of UPS genes reduces OPMD muscle defects suggesting a pathological increase of UPS activity in the disease. Quantification of proteasome activity confirms stronger activity in OPMD muscles, associated with degradation of myofibrillar proteins. Importantly, improvement of muscle structure and function in the presence of UPS mutants does not correlate with the levels of PABPN1 aggregation, but is linked to decreased degradation of muscle proteins. Oral treatment with the proteasome inhibitor MG132 is beneficial to the OPMD Drosophila model, improving muscle function although PABPN1 aggregation is enhanced. This functional study reveals the importance of increased UPS activity that underlies muscle atrophy in OPMD. It also provides a proof-of-concept that inhibitors of proteasome activity might be an attractive pharmacological approach for OPMD.