STK16

Transferrin-1 (Tsf1) is an extracellular insect protein with a high affinity for iron. The functions of Tsf1 are still poorly understood; however, Drosophila melanogaster Tsf1 has been shown to influence iron distribution in the fly body and to protect flies against some infections. The goal of this study was to better understand the physiological functions of Tsf1 in D. melanogaster by 1) investigating Tsf1 null phenotypes, 2) determining tissue-specific localization of Tsf1, 3) measuring the concentration of Tsf1 in hemolymph, 4) testing Tsf1 for bacteriostatic activity, and 5) evaluating the effect of metal and paraquat treatments on Tsf1 abundance. Flies lacking Tsf1 had more iron than wild-type flies in specialized midgut cells that take up iron from the diet; however, the absence of Tsf1 had no effect on the iron content of whole midguts, fat body, hemolymph, or heads. Thus, as previous studies have suggested, Tsf1 appears to have a minor role in iron transport. Tsf1 was abundant in hemolymph from larvae (0.4 μM), pupae (1.4 μM), adult females (4.4 μM) and adult males (22 μM). Apo-Tsf1 at 1 μM had bacteriostatic activity whereas holo-Tsf1 did not, suggesting that Tsf1 can inhibit microbial growth by sequestering iron in hemolymph and other extracellular environments. This hypothesis was supported by detection of secreted Tsf1 in tracheae, testes and seminal vesicles. Colocalization of Tsf1 with an endosome marker in oocytes suggested that Tsf1 may provide iron to developing eggs; however, eggs from mothers lacking Tsf1 had the same amount of iron as control eggs, and they hatched at a wild-type rate. Thus, the primary function of Tsf1 uptake by oocytes may be to defend against infection rather than to provide eggs with iron. In beetles, Tsf1 plays a role in protection against oxidative stress. In contrast, we found that flies lacking Tsf1 had a typical life span and greater resistance to paraquat-induced oxidative stress. In addition, Tsf1 abundance remained unchanged in response to ingestion of iron, cadmium or paraquat or to injection of iron. These results suggest that Tsf1 has a limited role in protection against oxidative stress in D. melanogaster.

The coronavirus disease 2019 (COVID-19) pandemic caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) continues to pose serious threats to global health. We previously reported that AAK1, BIKE and GAK, members of the Numb-associated kinase family, control intracellular trafficking of multiple RNA viruses during viral entry and assembly/egress. Here, using both genetic and pharmacological approaches, we probe the functional relevance of NAKs for SARS-CoV-2 infection. siRNA-mediated depletion of AAK1, BIKE, GAK, and STK16, the fourth member of the NAK family, suppressed SARS-CoV-2 infection in human lung epithelial cells. Both known and novel small molecules with potent AAK1/BIKE, GAK or STK16 activity suppressed SARS-CoV-2 infection. Moreover, combination treatment with the approved anti-cancer drugs, sunitinib and erlotinib, with potent anti-AAK1/BIKE and GAK activity, respectively, demonstrated synergistic effect against SARS-CoV-2 infection in vitro. Time-of-addition experiments revealed that pharmacological inhibition of AAK1 and BIKE suppressed viral entry as well as late stages of the SARS-CoV-2 life cycle. Lastly, suppression of NAKs expression by siRNAs inhibited entry of both wild type and SARS-CoV-2 pseudovirus. These findings provide insight into the roles of NAKs in SARS-CoV-2 infection and establish a proof-of-principle that pharmacological inhibition of NAKs can be potentially used as a host-targeted approach to treat SARS-CoV-2 with potential implications to other coronaviruses.

Serine/threonine kinase 16 (STK16) is crucial in on regulating tumor cell proliferation, apoptosis, and prognosis. Activated M1 macrophages regulate lung adenocarcinoma (LUAD) growth by releasing exosomes. This study aims to investigate the role of STK16 and then focus on the possible mechanisms through which exosomes derived from M1 macrophages play their roles in LUAD cells by targeting STK16. Clinical LUAD samples were used to evaluate the expression of STK16 and its association with prognosis. Exosomes were isolated from M0 and M1 macrophages by ultracentrifugation and were then identified by electron microscopy and western blotting. In vitro gain- and loss-of-function experiments with LUAD cells were performed to elucidate the functions of miR-181a-5p, ETS1, and STK16, and mouse xenograft models were used to verify the function of STK16 in vivo. Western blotting, quantitative real-time PCR, CCK-8 assay, cell apoptosis, immunohistochemistry staining, luciferase assay, ChIP assay, and bioinformatics analysis were performed to reveal the underlying mechanisms. High expression of STK16 was observed in LUAD tissues and cells, and higher expression of STK16 was associated with worse prognosis. Silencing STK16 expression inhibited cell proliferation and promoted apoptosis via the AKT1 pathway. Exosomes from M1 macrophages inhibited viability and promoted apoptosis by inhibiting STK16. Moreover, miR-181a-5p is the functional molecule in M1 macrophage-derived exosomes and plays a vital role in inhibiting cell proliferation and promoting apoptosis by targeting ETS1 and STK16. Hence, exosomes derived from M1 macrophages were capable of inhibiting viability and promoting apoptosis in LUAD via the miR-181a-5p/ETS1/STK16 axis.