The WD40 domain is one of the most abundant domains and is among the top interacting domains in eukaryotic genomes. The WD40 domain of ATG16L1 is essential for LC3 recruitment to endolysosomal membranes during non-canonical autophagy, but dispensable for canonical autophagy. Canonical autophagy was utilized by FMDV, while the relationship between FMDV and non-canonical autophagy is still elusive. In the present study, WD40 knockout (KO) PK15 cells were successfully generated via CRISPR/cas9 technology as a tool for studying the effect of non-canonical autophagy on FMDV replication. The results of growth curve analysis, morphological observation and karyotype analysis showed that the WD40 knockout cell line was stable in terms of growth and morphological characteristics. After infection with FMDV, the expression of viral protein, viral titers, and the number of copies of viral RNA in the WD40-KO cells were significantly greater than those in the wild-type PK15 cells. Moreover, RNA‒seq technology was used to sequence WD40-KO cells and wild-type cells infected or uninfected with FMDV. Differentially expressed factors such as Mx1, RSAD2, IFIT1, IRF9, IFITM3, GBP1, CXCL8, CCL5, TNFRSF17 were significantly enriched in the autophagy, NOD-like receptor signaling pathway, RIG-I-like receptor signaling pathway, Toll-like receptor signaling pathway, cytokine-cytokine receptor interaction and TNF signaling pathway, etc. The expression levels of differentially expressed genes were detected via qRT‒PCR, which was consistent with the RNA‒seq data. Here, we experimentally demonstrate for the first time that knockout of the WD40 domain of ATG16L1 enhances FMDV replication by downregulation innate immune factors. In addition, this result also indicates non-canonical autophagy inhibits FMDV replication. In total, our results play an essential role in regulating the replication level of FMDV and providing new insights into virus-host interactions and potential antiviral strategies.

Canonical autophagy is an evolutionarily conserved process that forms double-membrane structures and mediates the degradation of long-lived proteins (LLPs). Noncanonical autophagy (NCA) is an important alternative pathway involving the formation of microtubule-associated protein 1 light chain 3 (LC3)-positive structures that are independent of partial core autophagy proteins. NCA has been defined by the conjugation of ATG8s to single membranes (CASM). During canonical autophagy and NCA/CASM, LC3 undergoes a lipidation modification, and ATG16L1 is a crucial protein in this process. Previous studies have reported that the WDR domain of ATG16L1 is not necessary for canonical autophagy. However, our study found that WDR domain deficiency significantly impaired LLP degradation in basal conditions and slowed down LC3-II accumulation in canonical autophagy. We further demonstrated that the observed effect was due to a reduced interaction between ATG16L1 and FIP200/WIPI2, without affecting lysosome function or fusion. Furthermore, we also found that the WDR domain of ATG16L1 is crucial for chemical-induced NCA/CASM. The results showed that removing the WDR domain or introducing the K490A mutation in ATG16L1 significantly inhibited the NCA/CASM, which interrupted the V-ATPase-ATG16L1 axis. In conclusion, this study highlights the significance of the WDR domain of ATG16L1 for both canonical autophagy and NCA functions, improving our understanding of its role in autophagy.

Cytoplasmic dynein is a microtubule motor vital for cellular organization and division. It functions as a ~4-megadalton complex containing its cofactor dynactin and a cargo-specific coiled-coil adaptor. However, how dynein and dynactin recognize diverse adaptors, how they interact with each other during complex formation, and the role of critical regulators such as lissencephaly-1 (LIS1) protein (LIS1) remain unclear. In this study, we determined the cryo-electron microscopy structure of dynein-dynactin on microtubules with LIS1 and the lysosomal adaptor JIP3. This structure reveals the molecular basis of interactions occurring during dynein activation. We show how JIP3 activates dynein despite its atypical architecture. Unexpectedly, LIS1 binds dynactin\'s p150 subunit, tethering it along the length of dynein. Our data suggest that LIS1 and p150 constrain dynein-dynactin to ensure efficient complex formation.

WDR44 prevents ciliogenesis initiation by regulating RAB11-dependent vesicle trafficking. Here, we describe male patients with missense and nonsense variants within the WD40 repeats (WDR) of WDR44, an X-linked gene product, who display ciliopathy-related developmental phenotypes that we can model in zebrafish. The patient phenotypic spectrum includes developmental delay/intellectual disability, hypotonia, distinct craniofacial features and variable presence of brain, renal, cardiac and musculoskeletal abnormalities. We demonstrate that WDR44 variants associated with more severe disease impair ciliogenesis initiation and ciliary signaling. Because WDR44 negatively regulates ciliogenesis, it was surprising that pathogenic missense variants showed reduced abundance, which we link to misfolding of WDR autonomous repeats and degradation by the proteasome. We discover that disease severity correlates with increased RAB11 binding, which we propose drives ciliogenesis initiation dysregulation. Finally, we discover interdomain interactions between the WDR and NH2-terminal region that contains the RAB11 binding domain (RBD) and show patient variants disrupt this association. This study provides new insights into WDR44 WDR structure and characterizes a new syndrome that could result from impaired ciliogenesis.

BACKGROUND: The WD40 repeat (WDR) domain provides scaffolds for numerous protein-protein interactions in multiple biological processes. WDR domain 76 (WDR76) has complex functionality owing to its diversified interactions; however, its mechanism in LGG has not yet been reported.
METHODS: Transcriptomic data from public databases were multifariously analyzed to explore the role of WDR76 in LGG pathology and tumor immunity. Laboratory experiments were conducted to confirm these results.
RESULTS: The results first confirmed that high expression of WDR76 in LGG was not only positively associated with clinical and molecular features of malignant LGG, but also served as an independent prognostic factor that predicted shorter survival in patients with LGG. Furthermore, high expression of WDR76 resulted in the upregulation of oncogenes, such as PRC1 and NUSAP1, and the activation of oncogenic mechanisms, such as the cell cycle and Notch signaling pathway. Finally, WDR76 was shown to be involved in LGG tumor immunity by promoting the infiltration of immune cells, such as M2 macrophages, and the expression of immune checkpoints, such as PDCD1 (encoding PD-1).
CONCLUSIONS: This study shows for the first time the diagnostic and prognostic value of WDR76 in LGG and provides a novel personalized biomarker for future targeted therapy and immunotherapy. Thus, WDR76 may significantly improve the prognosis of patients with LGG.

OBJECTIVE: Over-activation of nuclear factor kappa B (NF-κB) was proven to be involved in the pathogenesis of preeclampsia. However, its regulation mechanism is not clear yet. This paper explored the role of WD repeat domain 5 (WDR5) in the development of late-onset preeclampsia and its relationship with NF-κB.
METHODS: WDR5 expression was detected in normal placentas and placentas from late-onset preeclampsia patients. CCK-8 and colony formation assays were conducted to appraise the proliferative ability of trophoblast. Migration and invasion were observed by wound healing and transwell assays. The interaction between WDR5 and NF-κB inhibitor I-kappa-B-alpha (IkBa) was verified by Co-immunoprecipitation analysis. Immunofluorescence was used to analyze the activation of NF-κB. Finally, we tested the role of WDR5 using the mice late-onset preeclampsia model.
RESULTS: WDR5 was highly expressed in the placentas of late-onset preeclampsia patients. WDR5 overexpression suppressed cell proliferation, migration, and invasion in trophoblast. WDR5 could interact with IkBa to activate NF-κB. Knockdown of NF-κB counteracted the anti-proliferative and anti-metastatic effects of WDR5 overexpression in trophoblast. In-vivo studies suggested that targeting WDR5 combated late-onset preeclampsia development.
CONCLUSIONS: Our finding provides new insights into the role of WDR5 in late-onset preeclampsia development.

Non-alcoholic fatty liver disease (NAFLD), characterized by hepatic steatosis, is one of the commonest causes of liver dysfunction. Adipose triglyceride lipase (ATGL) is closely related to lipid turnover and hepatic steatosis as the speed-limited triacylglycerol lipase in liver lipolysis. However, the expression and regulation of ATGL in NAFLD remain unclear. Herein, our results showed that ATGL protein levels were decreased in the liver tissues of high-fat diet (HFD)-fed mice, naturally obese mice, and cholangioma/hepatic carcinoma patients with hepatic steatosis, as well as in the oleic acid-induced hepatic steatosis cell model, while ATGL mRNA levels were not changed. ATGL protein was mainly degraded through the proteasome pathway in hepatocytes. Beta-transducin repeat containing (BTRC) was upregulated and negatively correlated with the decreased ATGL level in these hepatic steatosis models. Consequently, BTRC was identified as the E3 ligase for ATGL through predominant ubiquitination at the lysine 135 residue. Moreover, adenovirus-mediated knockdown of BTRC ameliorated steatosis in HFD-fed mouse livers and oleic acid-treated liver cells via upregulating the ATGL level. Taken together, BTRC plays a crucial role in hepatic steatosis as a new ATGL E3 ligase and may serve as a potential therapeutic target for treating NAFLD.

SUPPRESSOR OF MAX2-LIKE 6, 7, and 8 (SMXL6,7,8) function as repressors and transcription factors of the strigolactone (SL) signaling pathway, playing an important role in the development and stress tolerance in Arabidopsis thaliana. However, the molecular mechanism by which SMXL6,7,8 negatively regulate drought tolerance and ABA response remains largely unexplored. In the present study, the interacting protein and downstream target genes of SMXL6,7,8 were investigated. Our results showed that the substrate receptor for the CUL4-based E3 ligase DDB1-BINDING WD-REPEAT DOMAIN (DWD) HYPERSENSITIVE TO ABA DEFICIENT 1 (ABA1) (DWA1) physically interacted with SMXL6,7,8. The degradation of SMXL6,7,8 proteins were partially dependent on DWA1. Disruption of SMXL6,7,8 resulted in increased drought tolerance and could restore the drought-sensitive phenotype of the dwa1 mutant. In addition, SMXL6,7,8 could directly bind to the promoter of SUCROSE NONFERMENTING 1 (SNF1)-RELATED PROTEIN KINASE 2.3 (SnRK2.3) to repress its transcription. The mutations in SnRK2.2/2.3 significantly suppressed the hypersensitivity of smxl6/7/8 to ABA-mediated inhibition of seed germination. Conclusively, SMXL6,7,8 interact with DWA1 to negatively regulate drought tolerance and target ABA-response genes. These data provide insights into drought tolerance and ABA response in Arabidopsis via the SMXL6,7,8-mediated SL signaling pathway.

Heading date (flowering time), which greatly influences regional and seasonal adaptability in rice (Oryza sativa), is regulated by many genes in different photoperiod pathways. Here, we characterized a heading date gene, Early heading date 5 (Ehd5), using a modified bulked segregant analysis method. The ehd5 mutant showed late flowering under both short-day and long-day conditions, as well as reduced yield, compared to the wild type. Ehd5, which encodes a WD40 domain-containing protein, is induced by light and follows a circadian rhythm expression pattern. Transcriptome analysis revealed that Ehd5 acts upstream of the flowering genes Early heading date 1 (Ehd1), RICE FLOWERING LOCUS T 1 (RFT1), and Heading date 3a (Hd3a). Functional analysis showed that Ehd5 directly interacts with Rice outermost cell-specific gene 4 (Roc4) and Grain number, plant height, and heading date 8 (Ghd8), which might affect the formation of Ghd7-Ghd8 complexes, resulting in increased expression of Ehd1, Hd3a, and RFT1. In a nutshell, these results demonstrate that Ehd5 functions as a positive regulator of rice flowering and provide insight into the molecular mechanisms underlying heading date.

WD repeat and HMG-box DNA binding protein 1 (WDHD1) is a highly conserved gene from yeast to humans. It actively participates in DNA replication, playing a crucial role in DNA damage repair and the cell cycle, contributing to centromere formation and sister chromosome segregation. Notably, several studies have implicated WDHD1 in the development and progression of diverse tumor types, including esophageal carcinoma, pulmonary carcinoma, and breast carcinoma. Additionally, the inhibitor of WDHD1 has been found to enhance radiation sensitivity, improve drug resistance, and significantly decrease tumor cell proliferation. This comprehensive review aims to provide an overview of the molecular structure, biological functions, and regulatory mechanisms of WDHD1 in tumors, thereby establishing a foundation for future investigations and potential clinical applications of WDHD1.