KIFC1 is a motor protein of the Kinesin family and it is involved in spindle apparatus assembly, chromosome arrangement, and microfilament-mediated biological processes in mitosis. However, the specific function of KIFC1 in pig oocytes remains unclear. Here, in order to explore the function of KIFC1 in porcine oocytes, the AZ82 inhibitor was used to inhibit the activity of KIFC1. Our results showed when KIFC1 was inhibited, the polar body extrusion rate was obviously decreased, indicating that KIFC1 plays a crucial role in porcine oocytes. We next measured the spindle structure and chromosome arrangement via immunofluorescent staining and found both the rates of abnormal spindle and chromosome disorder increased significantly. By further analyzing the causes of the abnormal spindle, we found the acetylation of tubulin was disrupted. In addition, we also found the spindle position was impaired after KIFC1 inhibition, declaring the spindle migration was affected. Further analysis found cortex actin decreased and cytoplasmic actin increased after KIFC1 inhibition. In summary, we found that KIFC1 played a critical role in porcine oocytes maturation by controlling spindle apparatus via mediating the acetylation of microtubule and regulating the spindle migration via affecting actin dynamics.

The development of resistance remains one of the biggest challenges in clinical cancer patient care and it comprises all treatment modalities from chemotherapy to targeted or immune therapy. In solid malignancies, drug resistance is the result of adaptive processes occurring in cancer cells or the surrounding tumor microenvironment (TME). Future therapy attempts will therefore benefit from targeting both, tumor and stroma compartments and drug targets which affect both sides will be highly appreciated. In this review, we describe a seemingly paradoxical oncogenic mediator with this potential: The dual-specificity tyrosine-phosphorylation regulated kinase 1B (DYRK1B). DYRK1B promotes proliferative quiescence and yet is overexpressed or amplified in many hyperproliferative malignancies including ovarian cancer and pancreatic cancer. In particular the latter disease is a paradigmatic example for a therapy-recalcitrant and highly stroma-rich cancer entity. Here, recent evidence suggests that DYRK1B exerts its oncogenic features by installing a protective niche for cancer cells by directly affecting cancer cells but also the TME. Specifically, DYRK1B not only fosters cell-intrinsic processes like cell survival, chemoresistance, and disease recurrence, but it also upregulates TME and cancer cell-protective innate immune checkpoints and down-modulates anti-tumoral macrophage functionality. In this article, we outline the well-established cell-autonomous roles of DYRK1B and extend its importance to the TME and the control of the tumor immune stroma. In summary, DYRK1B appears as a single novel key player creating a safe haven for cancer cells by acting cell-intrinsically and-extrinsically, leading to the promotion of cancer cell survival, chemoresistance, and relapse. Thus, DYRK1B appears as an attractive drug target for future therapeutic approaches.

During the last years, there has been an increased effort in the discovery of selective and potent kinase inhibitors for targeted cancer therapy. Kinase inhibitors exhibit less toxicity compared to conventional chemotherapy, and several have entered the market. Mirk/Dyrk1B kinase is a promising pharmacological target in cancer since it is overexpressed in many tumors, and its overexpression is correlated with patients' poor prognosis. Mirk/Dyrk1B acts as a negative cell cycle regulator, maintaining the survival of quiescent cancer cells and conferring their resistance to chemotherapies. Many studies have demonstrated the valuable therapeutic effect of Mirk/Dyrk1B inhibitors in cancer cell lines, mouse xenografts, and patient-derived 3D-organoids, providing a perspective for entering clinical trials. Since the majority of Mirk/Dyrk1B inhibitors target the highly conserved ATP-binding site, they exhibit off-target effects with other kinases, especially with the highly similar Dyrk1A. In this review, apart from summarizing the data establishing Dyrk1B as a therapeutic target in cancer, we highlight the most potent Mirk/Dyrk1B inhibitors recently reported. We also discuss the limitations and perspectives for the structure-based design of Mirk/Dyrk1B potent and highly selective inhibitors based on the accumulated structural data of Dyrk1A and the recent crystal structure of Dyrk1B with AZ191 inhibitor.

Endogenous electric fields (EFs) have been demonstrated to facilitate wound healing by directing the migration of epidermal cells. Despite the identification of numerous molecules and signaling pathways that are crucial for the directional migration of keratinocytes under EFs, the underlying molecular mechanisms remain undefined. Previous studies have indicated that microtubule (MT) acetylation is linked to cell migration, while Paxillin exerts a significant influence on cell motility. Therefore, we postulated that Paxillin could enhance EF-induced directional migration of keratinocytes by modulating MT acetylation. In the present study, we observed that EFs (200 mV/mm) induced migration of human immortalized epidermal cells (HaCaT) towards the anode, while upregulating Paxillin, downregulating HDAC6, and increasing the level of microtubule acetylation. Our findings suggested that Paxillin plays a pivotal role in inhibiting HDAC6-mediated microtubule acetylation during directional migration under EF regulation. Conversely, downregulation of Paxillin decreased microtubule acetylation and electrotaxis of epidermal cells by promoting HDAC6 expression, and this effect could be reversed by the addition of tubacin, an HDAC6-specific inhibitor. Furthermore, we observed that EFs also mediated the polarization of Paxillin and acetylated α-tubulin, which is critical for directional migration. In conclusion, our study revealed that MT acetylation in EF-guided keratinocyte migration is regulated by the Paxillin/HDAC6 signaling pathway, providing a novel theoretical foundation for the molecular mechanism of EF-guided directional migration of keratinocytes.

Histone deacetylase 6 (HDAC6) has been shown to play an important role in allergic inflammation. This study hypothesized that novel downstream targets of HDAC6 would mediate allergic inflammation. Experiments employing HDAC6 knock out C57BL/6 mice showed that HDAC6 mediated passive cutaneous anaphylaxis (PCA) and passive systemic anaphylaxis (PSA). Antigen stimulation increased expression of N-myc (MYCN) and CXCL3 in an HDAC6-dependent manner in the bone marrow-derived mast cells. MYCN and CXCL3 were necessary for both PCA and PSA. The role of early growth response 3 (EGR3) in the regulation of HDAC6 expression has been reported. ChIP assays showed EGR3 as a direct regulator of MYCN. miR-34a-5p was predicted to be a negative regulator of MYCN. Luciferase activity assays showed miR-34a-5p as a direct regulator of MYCN. miR-34a-5p mimic negatively regulated PCA and PSA. MYCN decreased miR-34a-5p expression in antigen-stimulated rat basophilic leukemia cells (RBL2H3). MYCN was shown to bind to the promoter sequence of CXCL3. In an IgE-independent manner, recombinant CXCL3 protein increased expression of HDAC6, MYCN, and β-hexosaminidase activity in RBL2H3 cells. Mouse recombinant CXCL3 protein enhanced the angiogenic potential of the culture medium of RBL2H3. CXCL3 was necessary for the enhanced angiogenic potential of the culture medium of antigen-stimulated RBL2H3. The culture medium of RBL2H3 was able to induce M2 macrophage polarization in a CXCL3-dependent manner. Recombinant CXCL3 protein also increased the expression of markers of M2 macrophage. Thus, the identification of the novel role of HDAC6-MYCN-CXCL3 axis can help better understand the pathogenesis of anaphylaxis.

Cross-talk between Mirk/Dyrk1B kinase and Sonic hedgehog (Shh)/Gli pathway affects physiology and pathology. Here, we reveal a novel role for Dyrk1B in regulating ventral progenitor and neuron subtypes in the embryonic chick spinal cord (SC) via the Shh pathway. Using in ovo gain-and-loss-of-function approaches at E2, we report that Dyrk1B affects the proliferation and differentiation of neuronal progenitors at E4 and impacts on apoptosis specifically in the motor neuron (MN) domain. Especially, Dyrk1B overexpression decreases the numbers of ventral progenitors, MNs, and V2a interneurons, while the pharmacological inhibition of endogenous Dyrk1B kinase activity by AZ191 administration increases the numbers of ventral progenitors and MNs. Mechanistically, Dyrk1B overexpression suppresses Shh, Gli2 and Gli3 mRNA levels, while conversely, Shh, Gli2 and Gli3 transcription is increased in the presence of Dyrk1B inhibitor AZ191 or Smoothened agonist SAG. Most importantly, in phenotype rescue experiments, SAG restores the Dyrk1B-mediated dysregulation of ventral progenitors. Further at E6, Dyrk1B affects selectively the medial lateral motor neuron column (LMCm), consistent with the expression of Shh in this region. Collectively, these observations reveal a novel regulatory function of Dyrk1B kinase in suppressing the Shh/Gli pathway and thus affecting ventral subtypes in the developing spinal cord. These data render Dyrk1B a possible therapeutic target for motor neuron diseases.

The inter-neuronal communication occurring in extensively branched neuronal cells is achieved primarily through the microtubule (MT)-mediated axonal transport system. This mechanistically regulated system delivers cargos (proteins, mRNAs and organelles such as mitochondria) back and forth from the soma to the synapse. Motor proteins like kinesins and dynein mechanistically regulate polarized anterograde (from the soma to the synapse) and retrograde (from the synapse to the soma) commute of the cargos, respectively. Proficient axonal transport of such cargos is achieved by altering the microtubule stability via post-translational modifications (PTMs) of α- and β-tubulin heterodimers, core components constructing the MTs. Occurring within the lumen of MTs, K40 acetylation of α-tubulin via α-tubulin acetyl transferase and its subsequent deacetylation by HDAC6 and SIRT2 are widely scrutinized PTMs that make the MTs highly flexible, which in turn promotes their lifespan. The movement of various motor proteins, including kinesin-1 (responsible for axonal mitochondrial commute), is enhanced by this PTM, and dyshomeostasis of neuronal MT acetylation has been observed in a variety of neurodegenerative conditions, including Alzheimer's disease and Parkinson's disease (PD). PD is the second most common neurodegenerative condition and is closely associated with impaired MT dynamics and deregulated tubulin acetylation levels. Although the relationship between status of MT acetylation and progression of PD pathogenesis has become a chicken-and-egg question, our review aims to provide insights into the MT-mediated axonal commute of mitochondria and dyshomeostasis of MT acetylation in PD. The enzymatic regulators of MT acetylation along with their synthetic modulators have also been briefly explored. Moving towards a tubulin-based therapy that enhances MT acetylation could serve as a disease-modifying treatment in neurological conditions that lack it.

Medulloblastoma (MB) is the most common malignant brain tumor in children. Transcriptional profiling has so far delineated four major MB subgroups of which one is driven by uncontrolled Hedgehog (Hh) signaling (SHH-MB). This pathway is amenable to drug targeting, yet clinically approved compounds exclusively target the transmembrane component Smoothened (SMO). Unfortunately, drug resistance against SMO inhibitors is encountered frequently, making the identification of novel Hh pathway components mandatory, which could serve as novel drug targets in the future. Here, we have used MB as a tool to delineate novel modulators of Hh signaling and have identified the Acidic Nuclear Phosphoprotein 32 (ANP32) family of proteins as novel regulators. The expression of all three family members (ANP32A, ANP32B, ANP32E) is increased in Hh-induced MB and their expression level is negatively associated with overall survival in SHH-MB patients. Mechanistically, we could find that ANP32 proteins function as positive modulators of mammalian Hh signaling upstream of GLI transcription factors. These findings add hitherto unknown regulators to the mammalian Hh signaling cascade and might spur future translational efforts to combat Hh-driven malignancies.

Hepatocellular carcinoma (HCC) is one of the common malignancy and lacks effective therapeutic targets. Here, we demonstrated that ectopic expression of trophinin-associated protein (TROAP) dramatically drove HCC cell growth assessed by foci formation in monolayer culture, colony formation in soft agar and orthotopic liver transplantation in nude mice. Inversely, silencing TROAP expression with short-hairpin RNA attenuated the malignant proliferation of HCC cells in vitro and in vivo. Next, mechanistic investigation revealed that TROAP directly bound to dual specificity tyrosine phosphorylation regulated kinase 1A/B (DYRK1A/B), resulting in the cytoplasmic retention of proteins DYRK1A/B and promoting cell cycle process via activation of Akt/GSK-3β signaling. Combination of cisplatin with an inhibitor of DYRK1 AZ191 effectively inhibited tumor growth in mouse model for HCC cells with high level of TROAP. Clinically, TROAP was significantly upregulated by miR-142-5p in HCC tissues, which predicted the poor survival of patients with HCC. Therefore, TROAP/DYRK1/Akt axis may be a promising therapeutic target and prognostic indicator for patients with HCC.

Dual-specificity tyrosine phosphorylation-regulated kinase 1B (DYRK1B), also known as minibrain-related kinase (MIRK) is one of the best functionally studied members of the DYRK kinase family. DYRKs comprise a family of protein kinases that are emerging modulators of signal transduction pathways, cell proliferation and differentiation, survival, and cell motility. DYRKs were found to participate in several signaling pathways critical for development and cell homeostasis. In this review, we focus on the DYRK1B protein kinase from a functional point of view concerning the signaling pathways through which DYRK1B exerts its cell type-dependent function in a positive or negative manner, in development and human diseases. In particular, we focus on the physiological role of DYRK1B in behavior of stem cells in myogenesis, adipogenesis, spermatogenesis and neurogenesis, as well as in its pathological implication in cancer and metabolic syndrome. Thus, understanding of the molecular mechanisms that regulate signaling pathways is of high importance. Recent studies have identified a close regulatory connection between DYRK1B and the hedgehog (HH) signaling pathway. Here, we aim to bring together what is known about the functional integration and cross-talk between DYRK1B and several signaling pathways, such as HH, RAS and PI3K/mTOR/AKT, as well as how this might affect cellular and molecular processes in development, physiology, and pathology. Thus, this review summarizes the major known functions of DYRK1B kinase, as well as the mechanisms by which DYRK1B exerts its functions in development and human diseases focusing on the homeostasis of stem and cancer stem cells.