Mical1 deletion in tyrosinase expressing cells affects mouse running gaits.

Neuronal development is a highly regulated process that is dependent on the correct coordination of cellular responses to extracellular cues. In response to semaphorin axon guidance proteins, the MICAL1 protein is stimulated to produce reactive oxygen species that oxidize actin on specific methionine residues, leading to filamentous actin depolymerization and consequent changes in neuronal growth cone dynamics. Crossing genetically modified mice homozygous for floxed Mical1 (Mical1fl/fl) alleles with transgenic mice expressing Cre recombinase under the control of a tyrosinase gene enhancer/promoter (Tyr::Cre) enabled conditional Mical1 deletion. Immunohistochemical analysis showed Mical1 expression in the cerebellum, which plays a prominent role in the coordination of motor movements, with reduced Mical1 expression in Mical1fl/fl mice co-expressing Tyr::Cre. Analysis of the gaits of mice running on a treadmill showed that both male and female Mical1fl/fl, Tyr::Cre mutant mice had significant alterations to their striding patterns relative to wild-type mice, although the specific aspects of their altered gaits differed between the sexes. Additional motor tests that involved movement on a rotating rod, descending a vertical pole, or crossing a balance beam did not show significant differences between the genotypes, suggesting that the effect of the Mical1fl/fl, Tyr::Cre genetic modifications was only manifested during specific highly coordinated movements that contribute to running. These findings indicate that there is a behavioral consequence in Mical1fl/fl, Tyr::Cre mutant mice that affects motor control as manifested by alterations in their gait.

New targets and designed inhibitors of ASAP Arf-GAPs derived from structural characterization of the ASAP1/440-kD ankyrin-B interaction.

ASAP1 and its paralog ASAP2 belong to a PI4,5P2-dependent Arf GTPase-activating protein (Arf-GAP) family capable of modulating membrane and cytoskeletal dynamics. ASAPs regulate cell adhesive structures such as invadosomes and focal adhesions during cell attachment and migration. Malfunctioning of ASAP1 has been implicated in the malignant phenotypes of various cancers. Here, we discovered that the SH3 domain of ASAP1 or ASAP2 specifically binds to a 12-residue, positively charged peptide fragment from the 440 kDa giant ankyrin-B, a neuronal axon specific scaffold protein. The high-resolution structure of the ASAP1-SH3 domain in complex with the gAnkB peptide revealed a noncanonical SH3-ligand binding mode with high affinity and specificity. Structural analysis of the complex readily uncovered a consensus ASAP1-SH3 binding motif, which allowed the discovery of a number of previously unknown binding partners of ASAP1-SH3 including Clasp1/Clasp2, ALS2, ß-Pix, DAPK3, PHIP, and Limk1. Fittingly, these newly identified ASAP1 binding partners are primarily key modulators of the cytoskeletons. Finally, we designed a cell-penetrating, highly potent ASAP1 SH3 domain binding peptide with a Kd ∼7 nM as a tool for studying the roles of ASAPs in different cellular processes.

A hominoid-specific signaling axis regulating the tempo of synaptic maturation.

Human cortical neurons (hCNs) exhibit high dendritic complexity and synaptic density, and the maturation process is greatly protracted. However, the molecular mechanism governing these specific features remains unclear. Here, we report that the hominoid-specific gene TBC1D3 promotes dendritic arborization and protracts the pace of synaptogenesis. Ablation of TBC1D3 in induced hCNs causes reduction of dendritic growth and precocious synaptic maturation. Forced expression of TBC1D3 in the mouse cortex protracts synaptic maturation while increasing dendritic growth. Mechanistically, TBC1D3 functions via interaction with MICAL1, a monooxygenase that mediates oxidation of actin filament. At the early stage of differentiation, the TBC1D3/MICAL1 interaction in the cytosol promotes dendritic growth via F-actin oxidation and enhanced actin dynamics. At late stages, TBC1D3 escorts MICAL1 into the nucleus and downregulates the expression of genes related with synaptic maturation through interaction with the chromatin remodeling factor ATRX. Thus, this study delineates the molecular mechanisms underlying human neuron development.

MICAL1 promotes the proliferation in acute myeloid leukemia and is associated with clinical prognosis and immune infiltration.

Acute myeloid leukemia (AML) is one of the most common hematopoietic malignancies that has a poor prognosis and a high rate of relapse. Dysregulated metabolism plays an important role in AML progression. This study aimed to conduct a comprehensive analysis of MRGs using TCGA and GEO datasets and further explore the potential function of critical MRGs in AML progression. In this study, we identified 17 survival-related differentially expressed MRGs in AML using TCGA and GEO datasets. The 150 AML samples were divided into three molecular subtypes using 17 MRGs, and we found that three molecular subtypes exhibited a different association with ferroptosis, cuproptosis and m6A related genes. Moreover, a prognostic signature that comprised nine MRGs and had good predictive capacity was established by LASSO-Cox stepwise regression analysis. Among the 17 MRGs, our attention focused on MICAL1 which was highly expressed in many types of tumors, including AML and its overexpression was also confirmed in several AML cell lines. We also found that the expression of MICAL1 was associated with several immune cells. Moreover, functional experiments revealed that knockdown of MICAL1 distinctly suppressed the proliferation of AML cells. Overall, this study not only contributes to a deeper understanding of the molecular mechanisms underlying AML but also provides potential targets and prognostic markers for AML treatment. These findings offer robust support for further research into therapeutic strategies and mechanisms related to AML, with the potential to improve the prognosis and quality of life for AML patients. Nevertheless, further research is needed to validate these findings and explore more in-depth molecular mechanisms.

A novel splicing variant in MICAL-1 gene is associated with epilepsy.

Germline MICAL1 defects have been rarely reported in patients with epilepsy and the genotype-phenotype association remains unclear. In this study, the patient was a 4.6 years old girl who presented with onset of recurrent focal seizures with onset at age 3.4 years. EEG showed abnormal δ-wave activity in the right central and middle temporal lobe. Trio WES showed a novel heterozygous variant c.-43-1G > A in the MICAL1 gene in the patient and her normal mother. Minigene verified two abnormal transcripts due to the mutation, which was predicted to interrupt 5'UTR structures of MICAL1. The patient was clinically diagnosed with benign childhood epilepsy with centrotemporal spike (BECTS). As far as we know, this is the first BECTS case with documented MICAL1 mutation. Novel MICAL1 variant c.-43-1G > A putatively interrupted MICAL1 translation by changing 5'UTR structures and, however, further functioning study is needed.

A de novo pathogenic variant in MICAL-1 causes epilepsy with auditory features.

Familial epilepsy with auditory features (FEAF), previously known as autosomal-dominant lateral temporal lobe epilepsy (ADLTE) is a genetically heterogeneous syndrome, clinically characterized by focal seizures with prominent auditory symptoms. It is inherited with autosomal-dominant pattern with reduced penetrance (about 70%). Sporadic epilepsy with auditory features cases are more frequent and clinically indistinguishable from familial cases. One causal gene, MICAL-1, encodes MICAL-1, an intracellular multi-domain enzyme that is an important regulator of filamentous actin (F-actin) structures. Pathogenic variants in MICAL-1 account for approximately 7% of FEAF families. Here, we describe a de novo MICAL-1 pathogenic variant, p.Arg915Cys, in a sporadic case, an affected 21-year-old Italian man with no family history of epilepsy. Genetic testing was performed in the patient and his parents, using a next-generation sequencing panel. In cell-based assay, this variant significantly increased MICAL-1 oxidoreductase activity, which likely resulted in dysregulation of F-actin organization. This finding provides further support for a gain-of-function effect underlying MICAL-1-mediated epilepsy pathogenesis, as previously seen with other pathogenic variants. Furthermore, the case study provides evidence that de novo MICAL-1 pathogenic variants can occur in sporadic cases with epilepsy with auditory feature (EAF). PLAIN LANGUAGE SUMMARY: In this study, we report a new MICAL-1 pathogenic variant in a patient without family history for epilepsy, not inherited from his parents. MICAL-1 is a protein with enzymatic activity that reorganizes the structure of the cell. We proved the pathological effect of this variant by testing its enzymatic activity and found an increase of this activity. This result suggests that non-familial cases should be tested to find novel pathogenic variants in this gene.

PlexinA1 promotes gastric cancer migration through preventing MICAL1 protein ubiquitin/proteasome-mediated degradation in a Rac1-dependent manner.

Metastasis promotes the development of tumors and is a significant cause of gastric cancer death. For metastasis to proceed, tumor cells must become mobile by modulating their cytoskeleton. MICAL1 (Molecule Interacting with CasL1) is known as an actin cytoskeleton regulator, but the mechanisms by which it drives gastric cancer cell migration are still unclear. Analysis of gastric cancer tissues revealed that MICAL1 expression is dramatically upregulated in stomach adenocarcinoma (STAD) samples as compared to noncancerous stomach tissues. Patients with high MICAL1 expression had shorter overall survival (OS), post-progression survival (PPS) and first-progression survival (FPS) compared with patients with low MICAL1 expression. RNAi-mediated silencing of MICAL1 inhibited the expression of Vimentin, a protein involved in epithelial-mesenchymal transition. This effect correlates with a significant reduction in gastric cancer cell migration. MICAL1 overexpression reversed these preventive effects. Immunoprecipitation experiments and immunofluorescence assays revealed that PlexinA1 forms a complex with MICAL1. Importantly, specific inhibition of PlexinA1 blocked the Rac1 activation and ROS production, which, in turn, impaired MICAL1 protein stability by accelerating MICAL1 ubiquitin/proteasome-dependent degradation. Overexpression of PlexinA1 enhanced Rac1 activation, ROS production, MICAL1 and Vimentin expressions, and favored cell migration. In conclusion, this study identified MICAL1 as an important facilitator of gastric cancer cell migration, at least in part, by affecting Vimentin expression and PlexinA1 promotes gastric cancer cell migration by binding to and suppressing MICAL1 degradation in a Rac1/ROS-dependent manner.

Elucidating the role of MICAL1 in pan-cancer using integrated bioinformatics and experimental approaches.

Molecule interacting with CasL 1 (MICAL1) is a crucial protein involved in cell motility, axon guidance, cytoskeletal dynamics, and gene transcription. This pan-cancer study analyzed MICAL1 across 33 cancer types using bioinformatics and experiments. Dysregulated expression, diagnostic potential, and prognostic value were assessed. Associations with tumor characteristics, immune factors, and drug sensitivity were explored. Enrichment analysis revealed MICAL1's involvement in metastasis, angiogenesis, metabolism, and immune pathways. Functional experiments demonstrated its impact on renal carcinoma cells. These findings position MICAL1 as a potential biomarker and therapeutic target in specific cancers, warranting further investigation into its role in cancer pathogenesis.

Disassembly of bundled F-actin and cellular remodeling via an interplay of Mical, cofilin, and F-actin crosslinkers.

Cellular form and function are controlled by the assembly and stability of actin cytoskeletal structures-but disassembling/pruning these structures is equally essential for the plasticity and remodeling that underlie behavioral adaptations. Importantly, the mechanisms of actin assembly have been well-defined-including that it is driven by actin's polymerization into filaments (F-actin) and then often bundling by crosslinking proteins into stable higher-order structures. In contrast, it remains less clear how these stable bundled F-actin structures are rapidly disassembled. We now uncover mechanisms that rapidly and extensively disassemble bundled F-actin. Using biochemical, structural, and imaging assays with purified proteins, we show that F-actin bundled with one of the most prominent crosslinkers, fascin, is extensively disassembled by Mical, the F-actin disassembly enzyme. Furthermore, the product of this Mical effect, Mical-oxidized actin, is poorly bundled by fascin, thereby further amplifying Mical's disassembly effects on bundled F-actin. Moreover, another critical F-actin regulator, cofilin, also affects fascin-bundled filaments, but we find herein that it synergizes with Mical to dramatically amplify its disassembly of bundled F-actin compared to the sum of their individual effects. Genetic and high-resolution cellular assays reveal that Mical also counteracts crosslinking proteins/bundled F-actin in vivo to control cellular extension, axon guidance, and Semaphorin/Plexin cell-cell repulsion. Yet, our results also support the idea that fascin-bundling serves to dampen Mical's F-actin disassembly in vitro and in vivo-and that physiologically relevant cellular remodeling requires a fine-tuned interplay between the factors that build bundled F-actin networks and those that disassemble them.

MICAL-mediated oxidation of actin and its effects on cytoskeletal and cellular dynamics.

Actin and its dynamic structural remodelings are involved in multiple cellular functions, including maintaining cell shape and integrity, cytokinesis, motility, navigation, and muscle contraction. Many actin-binding proteins regulate the cytoskeleton to facilitate these functions. Recently, actin's post-translational modifications (PTMs) and their importance to actin functions have gained increasing recognition. The MICAL family of proteins has emerged as important actin regulatory oxidation-reduction (Redox) enzymes, influencing actin's properties both in vitro and in vivo. MICALs specifically bind to actin filaments and selectively oxidize actin's methionine residues 44 and 47, which perturbs filaments' structure and leads to their disassembly. This review provides an overview of the MICALs and the impact of MICAL-mediated oxidation on actin's properties, including its assembly and disassembly, effects on other actin-binding proteins, and on cells and tissue systems.

Crystal Structure of the SH3 Domain of ASAP1 in Complex with the Proline Rich Motif (PRM) of MICAL1 Reveals a Unique SH3/PRM Interaction Mode.

SH3 domains are common protein binding modules. The target sequence of SH3 domains is usually a proline-rich motif (PRM) containing a minimal "PxxP" sequence. The mechanism of how different SH3 domains specifically choose their targets from vast PxxP-containing sequences is still not very clear, as many reported SH3/PRM interactions are weak and promiscuous. Here, we identified the binding of the SH3 domain of ASAP1 to the PRM of MICAL1 with a sub-µM binding affinity, and determined the crystal structure of ASAP1-SH3 and MICAL1-PRM complex. Our structural and biochemical analyses revealed that the target-binding pocket of ASAP1-SH3 contains two negatively charged patches to recognize the "xPx + Px+" sequence in MICAL1-PRM and consequently strengthen the interaction, differing from the typical SH3/PRM interaction. This unique PRM-binding pocket is also found in the SH3 domains of GTPase Regulator associated with focal adhesion kinase (GRAF) and Src kinase associated phosphoprotein 1 (SKAP1), which we named SH3AGS. In addition, we searched the Swiss-Prot database and found ~130 proteins with the SH3AGS-binding PRM in silico. Finally, gene ontology analysis suggests that the strong interaction between the SH3AGS-containing proteins and their targets may play roles in actin cytoskeleton regulation and vesicle trafficking.

PlexinA1-deficient mice exhibit decreased cell density and augmented oxidative stress in parvalbumin-expressing interneurons in the medial prefrontal cortex.

PlexinA1 (PlxnA1) is a transmembrane receptor for semaphorins (Semas), a large family of axonal guidance cues vital during neural development. PlxnA1 is expressed in embryonic interneurons, and PlxnA1 deletion in mice leads to less interneurons in the developing cortex. In addition, PlxnA1 has been identified as a schizophrenia susceptibility gene. In our previous study, PlxnA1 knockout (KO) mice under a BALB/cAJ genetic background exhibited significantly increased self-grooming and reduced prepulse inhibition, a reliable phenotype for investigating the neurobiology of schizophrenia. However, the mechanism underlying the abnormal behavior of PlxnA1 KO mice remains unclear. We first confirmed PlxnA1 mRNA expression in parvalbumin-expressing interneurons (PV cells) in the medial prefrontal cortex (mPFC) of adult mice. Immunohistochemical analysis (IHC) showed significantly decreased densities of both GABAergic neurons and PV cells in the mPFC of PlxnA1 KO mice compared with wild type mice (WT). PV cells were found to express molecule interacting with CasL 1 (MICAL1), an effector involved in Sema-Plxn signaling for axon guidance, suggesting MICAL1 and PlxnA1 co-expression in PV cells. Furthermore, IHC analysis of 8-oxo-dG, an oxidative stress marker, revealed significantly increased oxidative stress in PlxnA1-deficient PV cells compared with WT. Thus, increased oxidative stress and decreased PV cell density in the mPFC may determine the onset of PlxnA1 KO mice's abnormal behavior. Accordingly, deficient PlxnA1-mediated signaling may increase oxidative stress in PV cells, thereby disrupting PV-cell networks in the mPFC and causing abnormal behavior related to neuropsychiatric diseases.

High MICAL1 expression correlates with cancer progression and immune infiltration in renal clear cell carcinoma.

BACKGROUND: Molecule interacting with CasL 1 (MICAL1), a multidomain flavoprotein monooxygenase, is strongly involved in the biological processes related to cancer cell proliferation and metastasis. However, there were few reports on the clinical significance of MICAL1 in renal clear cell carcinoma. METHODS: The expression and prognostic value of MICAL1 in renal clear cell carcinoma were explored using immunohistochemical assays, public TCGA-KIRC databases and multiple analysis methods, including survival analysis, univariate and multivariate analyses, KEGG and GSEA. Wound healing and Transwell assays were performed to check the 786-O cell and Caki-1 cell migration abilities after knockdown of MICAL1. Western blotting was used to assess the regulatory effect of MICAL1 on the Rac1 activation. Additionally, the function of MICAL1 and the correlations between MICAL1 and immune infiltration levels in KIRC were investigated using TIMER and TISIDB. RESULTS: MICAL1 expression was significantly higher in carcinoma tissue compared with non-cancerous tissue. A survival analysis revealed that patients with high MICAL1 expression had shorter overall survival (OS) and disease-specific survival (DSS) compared with patients with low MICAL1 expression. ROC analysis also confirmed that MICAL1 has a high diagnostic value in KIRC. Importantly, the univariate and multivariate Cox analysis further confirmed that high MICAL1 expression was an independent risk factor for OS in patients with KIRC. In accordance with this, knockdown of MICAL1 expression decreased Rac1 activation and cell migration. KEGG and GSEA analysis revealed that the immune infiltration and Ras signaling pathways were significantly upregulated in the high MICAL1 expression group. In terms of immune infiltrating levels, MICAL1 expression was positively associated with CD8+/Treg cell infiltration levels. Specifically, bioinformatic analysis showed that MICAL1 expression had strong relationships with various T cell exhaustion markers. CONCLUSIONS: MICAL1 expression may act as a prognostic biomarker for determining the prognosis in renal clear cell carcinoma and plays an important role in regulating tumor immune microenvironment and cell migratory capacity.

MICAL1 facilitates pancreatic cancer proliferation, migration, and invasion by activating WNT/ß-catenin pathway.

BACKGROUND: MICAL1 is involved in the malignant processes of several types of cancer; however, the role of MICAL1 in pancreatic cancer (PC) has not been well-characterized. This study aimed to investigate the expression and function of MICAL1 in PC. METHODS: RT-qPCR and immunohistochemistry were used to detect MICAL1 expression in PC and adjacent nontumor tissues. Cell Counting Kit-8, EdU, clone formation, wound healing, and Transwell assays as well as animal models were used to investigate the effects of overexpression or inhibition of MICAL1 expression on the proliferation, invasion, and metastasis of PC cells. RNA-seq was used to explore the main pathway underlying the functions of MICAL1. Proteomics, mass spectrometry, and co-immunoprecipitation assays were used to investigate the interaction of proteins with MICAL1. Rescue experiments were conducted to validate these findings. RESULTS: Both MICAL1 mRNA and protein levels were upregulated in PC tissues compared with matched adjacent nontumor tissues. The expression level of MICAL1 was associated with the proliferative and metastatic status of PC. Repression of MICAL1 significantly inhibited PC cell growth, migration, and invasion in vitro and in vivo. RNA sequencing analysis indicated that MICAL1 was closely correlated with the WNT pathway. Overexpression of MICAL1 (1) promoted the phosphorylation of TBC1D1 at the Ser660 site, (2) facilitated the distribution of FZD7 on the cytomembrane, (3) inhibited the degradation of FZD7 in the lysosome, and (4) activated the WNT pathway. CONCLUSIONS: MICAL1 was upregulated in PC and involved in stimulating the progression of PC cells by activating the WNT/ß-catenin signaling pathway. Therefore, MICAL1 is a potential therapeutic target for PC.

MICAL1 Monooxygenase in Autosomal Dominant Lateral Temporal Epilepsy: Role in Cytoskeletal Regulation and Relation to Cancer.

Autosomal dominant lateral temporal epilepsy (ADLTE) is a genetic focal epilepsy associated with mutations in the LGI1, RELN, and MICAL1 genes. A previous study linking ADLTE with two MICAL1 mutations that resulted in the substitution of a highly conserved glycine residue for serine (G150S) or a frameshift mutation that swapped the last three C-terminal amino acids for 59 extra residues (A1065fs) concluded that the mutations increased enzymatic activity and promoted cell contraction. The roles of the Molecule Interacting with CasL 1 (MICAL1) protein in tightly regulated semaphorin signaling pathways suggest that activating MICAL1 mutations could result in defects in axonal guidance during neuronal development. Further studies would help to illuminate the causal relationships of these point mutations with ADLTE. In this review, we discuss the proposed pathogenesis caused by mutations in these three genes, with a particular emphasis on the G150S point mutation discovered in MICAL1. We also consider whether these types of activating MICAL1 mutations could be linked to cancer.

MICAL1 inhibits colorectal cancer cell migration and proliferation by regulating the EGR1/ß-catenin signaling pathway.

MICAL1 has been reported to be involved in the malignant processes of several types of cancer cells, however, the roles of MICAL1 in colorectal cancer (CRC) have not been well-characterized. This study aims to investigate the cellular functions and molecular mechanisms of MICAL1 in CRC cells. Here, we found that both mRNA and protein levels of MICAL1 were down-regulated in colorectal cancer tissues compared with matched adjacent non-tumor tissues, and the expression level of MICAL1 was correlated with the metastatic status of colorectal cancer. Importantly, overexpression of MICAL1 significantly inhibited colorectal cancer cell migration and growth, and increased the level of E-cadherin and Occludin, and suppressed the expression level of Vimentin and N-cadherin; while silencing of MICAL1 promoted CRC cell migration and enhanced EMT. In addition, MICAL1 overexpression significantly inhibited the proliferation and growth of CRC in vitro and in vivo. Moreover, RNA sequencing and bioinformatics analysis identified that MICAL1 was closely correlated with "cell migration", "cell cycle" and "ß-catenin signaling" genesets. Mechanistically, overexpression of MICAL1 downregulated the mRNA level of EGR1 and ß-catenin, decreased the protein level and nuclear translocation of ß-catenin, and inhibited the transcriptions of ß-catenin downstream targets, c-myc and cyclin D1. The ectopic expression of EGR1 or ß-catenin can significantly block the MICAL1-mediated inhibitory effects. Collectively, MICAL1 is down-regulated in CRC, and plays an inhibitory role in the migration and growth of CRC cells by suppressing the ERG1/ß-catenin signaling pathway.

MICAL1 regulates actin cytoskeleton organization, directional cell migration and the growth of human breast cancer cells as orthotopic xenograft tumours.

The Molecule Interacting with CasL 1 (MICAL1) monooxygenase has emerged as an important regulator of cytoskeleton organization via actin oxidation. Although filamentous actin (F-actin) increases MICAL1 monooxygenase activity, hydrogen peroxide (H2O2) is also generated in the absence of F-actin, suggesting that diffusible H2O2 might have additional functions. MICAL1 gene disruption by CRISPR/Cas9 in MDA MB 231 human breast cancer cells knocked out (KO) protein expression, which affected F-actin organization, cell size and motility. Transcriptomic profiling revealed that MICAL1 deletion significantly affected the expression of over 700 genes, with the majority being reduced in their expression levels. In addition, the absolute magnitudes of reduced gene expression were significantly greater than the magnitudes of increased gene expression. Gene set enrichment analysis (GSEA) identified receptor regulator activity as the most significant negatively enriched molecular function gene set. The prominent influence exerted by MICAL1 on F-actin structures was also associated with changes in the expression of several serum-response factor (SRF) regulated genes in KO cells. Moreover, MICAL1 disruption attenuated breast cancer tumour growth in vivo. Elevated MICAL1 gene expression was observed in invasive breast cancer samples from human patients relative to normal tissue, while MICAL1 amplification or point mutations were associated with reduced progression free survival. Collectively, these results demonstrate that MICAL1 gene disruption altered cytoskeleton organization, cell morphology and migration, gene expression, and impaired tumour growth in an orthotopic in vivo breast cancer model, suggesting that pharmacological MICAL1 inhibition could have therapeutic benefits for cancer patients.

Enhanced Production of the Mical Redox Domain for Enzymology and F-actin Disassembly Assays.

To change their behaviors, cells require actin proteins to assemble together into long polymers/filaments-and so a critical goal is to understand the factors that control this actin filament (F-actin) assembly and stability. We have identified a family of unusual actin regulators, the MICALs, which are flavoprotein monooxygenase/hydroxylase enzymes that associate with flavin adenine dinucleotide (FAD) and use the co-enzyme nicotinamide adenine dinucleotide phosphate (NADPH) in Redox reactions. F-actin is a specific substrate for these MICAL Redox enzymes, which oxidize specific amino acids within actin to destabilize actin filaments. Furthermore, this MICAL-catalyzed reaction is reversed by another family of Redox enzymes (SelR/MsrB enzymes)-thereby revealing a reversible Redox signaling process and biochemical mechanism regulating actin dynamics. Interestingly, in addition to the MICALs' Redox enzymatic portion through which MICALs covalently modify and affect actin, MICALs have multiple other domains. Less is known about the roles of these other MICAL domains. Here we provide approaches for obtaining high levels of recombinant protein for the Redox only portion of Mical and demonstrate its catalytic and F-actin disassembly activity. These results provide a ground state for future work aimed at defining the role of the other domains of Mical - including characterizing their effects on Mical's Redox enzymatic and F-actin disassembly activity.

MICAL1 (molecule interacting with CasL 1) protects oligodendrocyte cells from oxidative injury through regulating apoptosis, autophagy in spinal cord injury.

Molecule's mechanism of action interacting with CasL 1 (MICAL1) in spinal cord injury (SCI) is unclear. This study aimed to detect the function of MICAL1 in SCI. Western blot was used to analyze the change of MICAL1 in vivo. Immunofluorescence staining was used to detect the location of MICAL1 expression. Oligodendrocyte cells were treated with H2O2 to induce oxidative injury. Subsequently, siRNA transfection was performed to decrease MICAL1 expression in oligodendrocyte cells. Then, the effects of MICAL1 on oxidative stress, apoptosis, and autophagy were assessed. We found that silencing of MICAL1 could significantly reduce the levels of the nuclear factor erythroid 2-related factor 2 (Nrf2), increase the expression of pro-apoptotic factors (Bax and C-caspase 3), decrease the levels of anti-apoptotic factor (Bcl-2) and pro-autophagy factors (Beclin1 and LC3B). Therefore, MICAL1 is a potential target gene for SCI clinical therapy.

Identification of novel TGF-ß regulated genes with pro-migratory roles.

Transforming growth factor-ß (TGF-ß) signaling plays fundamental roles in the development and homeostasis of somatic cells. Dysregulated TGF-ß signaling contributes to cancer progression and relapse to therapies by inducing epithelial-to-mesenchymal transition (EMT), enriching cancer stem cells, and promoting immunosuppression. Although many TGF-ß-regulated genes have been identified, only a few datasets were obtained by next-generation sequencing. In this study, we performed RNA-sequencing analysis of MCF10A cells and identified 1166 genes that were upregulated and 861 genes that were downregulated by TGF-ß. Gene set enrichment analysis revealed that focal adhesion and metabolic pathways were the top enriched pathways of the up- and downregulated genes, respectively. Genes in these pathways also possess significant predictive value for renal cancers. Moreover, we confirmed that TGF-ß induced expression of MICAL1 and 2, and the histone demethylase, KDM7A, and revealed their regulatory roles on TGF-ß-induced cell migration. We also show a critical effect of KDM7A in regulating the acetylation of H3K27 on TGF-ß-induced genes. In sum, this study identified novel effectors that mediate the pro-migratory role of TGF-ß signaling, paving the way for future studies that investigate the function of MICAL family members in cancer and the novel epigenetic mechanisms downstream TGF-ß signaling.