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Bone loss remains a major health concern for astronauts during space flight. Increased osteoclast activity is one of the main causes of bone loss in astronauts subjected to microgravity conditions. However, the underlying molecular mechanisms remain unclear. Microtubule actin crosslinking factor 1 (MACF1) has been implicated in the regulation of cytoskeletal distribution, cell migration, and cell differentiation. Our previous studies have shown that MACF1 promotes the differentiation of pre-osteoclasts. However, whether MACF1 regulates the migration and cytoskeletal arrangement of pre-osteoclasts under microgravity conditions has not yet been elucidated. In this study, we used a hind-limb unloading (HLU) mouse model and a random positioning machine (RPM) to simulate the effects of microgravity on pre-osteoclasts. In the HLU mouse model, the expression of MACF1 was upregulated in primary pre-osteoclasts of mice, accompanied by enhanced migration in vivo. Moreover, simulated microgravity using the RPM also promoted MACF1 expression and migration of pre-osteoclasts in vitro. Additionally, knockdown of MACF1 disrupted cytoskeletal arrangement (F-actin and microtubules) and further inhibited the migration of pre-osteoclasts via the RhoA/ROCK1 signaling pathway. We further demonstrated that knockdown of MACF1 disrupted the enhanced migration and cytoskeleton arrangement of pre-osteoclasts induced by simulated microgravity. These data demonstrate that MACF1 positively regulates the migration and cytoskeletal organization of pre-osteoclasts under simulated microgravity, suggesting that MACF1 may be a therapeutic target for the treatment of bone loss induced by microgravity. • MACF1 is a microgravity sensitive gene. • Simulated microgravity induced MACF1 expression and migration of pre-osteoclasts. • MACF1 knockdown attenuated the cytoskeleton arrangement of pre-osteoclasts. • MACF1 knockdown inhibited the migration via the RhoA/Rock1 pathway. • MACF1 knockdown prevented the migration and cytoskeleton arrangement of pre-osteoclasts induced by simulated microgravity. [ABSTRACT FROM AUTHOR]

Spectraplakins are a family of evolutionarily conserved gigantic proteins and play critical roles in many cytoskeleton-related processes. Microtubule actin crosslinking factor 1 (MACF1) is one of the most versatile spectraplakin with multiple isoforms. As a broadly expressed mammalian spectraplakin, MACF1 is important in maintaining normal functions of many tissues. The loss-of-function studies using knockout mouse models reveal the pivotal roles of MACF1 in embryo development, skin integrity maintenance, neural development, bone formation, and colonic paracellular permeability. Mutation in the human MACF1 gene causes a novel myopathy genetic disease. In addition, abnormal expression of MACF1 is associated with schizophrenia, Parkinson’s disease, cancer and osteoporosis. This demonstrates the crucial roles of MACF1 in physiology and pathology. Here, we review the research advances of MACF1’s roles in specific tissue and in human diseases, providing the perspectives of MACF1 for future studies. [ABSTRACT FROM AUTHOR]

Osteoclasts are responsible for bone resorption and play essential roles in causing bone diseases such as osteoporosis. Microtubule actin crosslinking factor 1 (MACF1) is a large spectraplakin protein that has been implicated in regulating cytoskeletal distribution, cell migration, cell survival and cell differentiation. However, whether MACF1 regulates the differentiation of osteoclasts has not been elucidated. In this study, we found that the expression of MACF1 was increased in primary bone marrow-derived monocytes (BMMs) of osteoporotic mice and was downregulated during receptor activator of nuclear factor kappa-B ligand (RANKL)-induced osteoclastogenesis of pre-osteoclast cell lines RAW264.7 cells. RAW264.7 cells were transfected with shMACF1 using a lentiviral vector to study the role of MACF1 in osteoclastogenic differentiation. Knockdown of MACF1 in RAW264.7 cells inhibited the formation of multinucleated osteoclasts and decreased the expression of osteoclast-marker genes (Ctsk , Acp5 , Mmp9 and Oscar) during RANKL-induced osteoclastogenesis. Additionally, knockdown of MACF1 disrupted actin ring formation in osteoclasts and further blocked the bone resorption activity of osteoclasts by reducing the area and depth of pits. Knockdown of MACF1 had no effect on the survival of pre-osteoclasts and mature osteoclasts. We further established that knockdown of MACF1 attenuated the phosphorylation of Akt and GSK3β and inhibited the expression of its downstream target NFATc1. Akt activator rescued the inhibition of osteoclast differentiation by MACF1 knockdown. These data demonstrate that MACF1 positively regulates osteoclast differentiation via the Akt/GSK3β/NFATc1 signalling pathway, suggesting that targeting MACF1 may be a novel therapeutic approach against osteoporosis. • MACF1 knockdown inhibited RANKL-induced osteoclast differentiation and activity. • MACF1 knockdown has no effect on the survival of pre-osteoclasts and mature osteoclasts. • MACF1 knockdown attenuated the phosphorylation of Akt and GSK3β induced by RANKL. • MACF1 knockdown attenuated RANKL-induced osteoclast differentiation by regulating the Akt/GSK3β/NFATc1 signalling pathway. [ABSTRACT FROM AUTHOR]