Hlavackova, K. | bioRxiv Science

bioRxiv2023

Systematic microRNA screening identifies miR-19b as regulator of temozolomide response through targeting PP2A serine-threonine phosphatase in glioblastoma

Despite the standard of care, glioblastoma IDH wildtype (GBM) inevitably recurs, underscoring the need to develop new treatment strategies. To address the role of microRNAs in temozolomide (TMZ) response, we performed functional microRNA screens and consistently identified miR-19b. Our study reveals a novel axis between miR-19b and PPP2R5E subunit of serine/threonine protein phosphatase PP2A and establishes a so far unappreciated contribution of miR-19b in TMZ resistance of GBM. Specifically, our results demonstrate that attenuation of miR-19b in GBM cell lines and glioblastoma stem cells (GSCs) induces DNA damage, which further enhances the cytotoxic effects of TMZ treatment. We confirmed TMZ resistance induced by knocking down PPP2R5E in orthotopic mouse xenografts of GSCs. Furthermore, our results indicate that treating cells with the PP2A-activating drug FTY720 or knocking down endogenous PP2A-inhibiting proteins potentiates the cytotoxic effects of TMZ. MiR-19b attenuation or PPP2R5E activation could potentially be exploited in adjuvant therapy of GBM patients.

cancer biology↗

bioRxiv2022

Genetic manipulation of stress-induced mitogen-activated protein kinase modulates early stages of the nodulation process in Medicago sativa

Leguminous plants have established a mutualistic endosymbiotic interaction with nitrogen-fixing rhizobia to secure nitrogen sources in new specialized organs called root nodules. Before nodule formation, the development of early symbiotic structures is essential for rhizobia docking, internalization, targeted delivery, and intracellular accommodation. We have recently reported that overexpression of stress-induced mitogen-activated protein kinase (SIMK) in alfalfa affects root hair, nodule and shoot formation. However, detailed subcellular spatial distribution, activation, and developmental relocation of SIMK during the early stages of alfalfa nodulation remain unclear. Here, we qualitatively and quantitatively characterized SIMK distribution patterns in rhizobium-infected root hairs using live-cell imaging and immunolocalization, employing alfalfa stable transgenic lines with genetically manipulated SIMK abundance and kinase activity. In the SIMKK-RNAi line, showing downregulation of SIMKK and SIMK, we found a considerably decreased accumulation of phosphorylated SIMK around infection pockets and infection threads, which was strongly increased in the GFP-SIMK line, constitutively overexpressing GFP-tagged SIMK. Thus, genetically manipulated SIMK modulates root hair capacity to form infection pockets and infection threads. These results shed new light on SIMK spatio-temporal participation in the early interactions between alfalfa and rhizobia, and its internalization into root hairs, showing that local accumulation of active SIMK indeed modulates nodulation in alfalfa. One sentence summaryGenetic manipulation of SIMK in alfalfa revealed that SIMK modulates root hair capacity to form infection pockets and infection threads during the early interactions between alfalfa and rhizobia.

plant biology↗