Dataset Information

A requirement for PAK1 to support mitochondrial function and maintain cellular redox balance via electron transport chain proteins to prevent β-cell apoptosis.

ABSTRACT:

Objective

p21 (Cdc42/Rac1) activated Kinase 1 (PAK1) is a candidate susceptibility factor for type 2 diabetes (T2D). PAK1 is depleted in the islets from T2D donors, compared to control individuals. In addition, whole-body PAK1 knock out (PAK1-KO) in mice worsens the T2D-like effects of high-fat diet. The current study tested the effects of modulating PAK1 levels only in β-cells.

Materials/methods

β-cell-specific inducible PAK1 KO (βPAK1-iKO) mice were generated and used with human β-cells and T2D islets to evaluate β-cell function.

Results

βPAK1-iKO mice exhibited glucose intolerance and elevated β-cell apoptosis, but without peripheral insulin resistance. β-cells from βPAK-iKO mice also contained fewer mitochondria per cell. At the cellular level, human PAK1-deficient β-cells showed blunted glucose-stimulated insulin secretion and reduced mitochondrial function. Mitochondria from human PAK1-deficient β-cells were deficient in the electron transport chain (ETC) subunits CI, CIII, and CIV; NDUFA12, a CI complex protein, was identified as a novel PAK1 binding partner, and was significantly reduced with PAK1 knockdown. PAK1 knockdown disrupted the NAD⁺/NADH and NADP⁺/NADPH ratios, and elevated ROS. An imbalance of the redox state due to mitochondrial dysfunction leads to ER stress in β-cells. PAK1 replenishment in the β-cells of T2D human islets ameliorated levels of ER stress markers.

Conclusions

These findings support a protective function for PAK1 in β-cells. The results support a new model whereby the PAK1 in the β-cell plays a required role upstream of mitochondrial function, via maintaining ETC protein levels and averting stress-induced β-cell apoptosis to retain healthy functional β-cell mass.

SUBMITTER: Ahn M

PROVIDER: S-EPMC8123936 | biostudies-literature |

REPOSITORIES: biostudies-literature

ACCESS DATA

Similar Datasets

Project description:Phosphorylation of the major photosynthetic light harvesting antenna proteins by STN7 kinase balances excitation between PSII and PSI. Phosphorylation of such abundant proteins is unique, differing distinctively from conventional tasks of protein kinases in phosphorylation of low abundance proteins in signaling cascades. Excitation balance between PSII and PSI is critical for redox homeostasis between the plastoquinone and plastocyanin pools and PSI electron acceptors, determining the capacity of the thylakoid membrane to produce reactive oxygen species (ROS) that operate as signals relaying information between chloroplasts and other cellular compartments. STN7 has also been proposed to be a conventional signaling kinase, instigating the phosphorylation cascade required for coordinated expression of photosynthesis genes and assembly of the photosynthetic machinery. The absence of STN7 kinase, however, does not prevent plants from sensing redox imbalance and adjusting the stoichiometry of the photosynthetic machinery to restore redox homeostasis. This suggests that STN7 is not essential for signaling between the chloroplast and the nucleus. Here we discuss the evolution and functions of the STN7 and other thylakoid protein kinases and phosphatases, and the inherent difficulties in analyzing signaling cascades initiated from the photosynthetic machinery. Based on our analyses of literature and publicly available expression data, we conclude that STN7 exerts it signaling effect primarily by controlling chloroplast ROS homeostasis through maintaining steady-state phosphorylation of the light harvesting II proteins and the redox balance in the thylakoid membrane. ROS are important signaling molecules with a direct effect on the development of jasmonate, which in turn relays information out from the chloroplast. We propose that thylakoid membrane redox homeostasis, regulated by SNT7, sends cell-wide signals that reprogram the entire hormonal network in the cell.

Project description:To investigate if and how the integrity of the mitochondrial electron transport chain (mETC) influences susceptibility of Arabidopsis toward Colletotrichum higginsianum, we have selected previously characterized mutants with defects at different stages of the mETC, namely, the complex I mutant ndufs4, the complex II mutant sdh2-1, the complex III mutant ucr8-1, and a mutant of the uncoupling protein ucp1-2. Relative to wild type, the selected complex I, II, and III mutants showed decreased total respiration, increased alternative respiration, as well as increased redox charge of the NADP(H) pool and decreased redox charge of the NAD(H) pool in the dark. In the light, mETC mutants accumulated free amino acids, albeit to varying degrees. Glycine and serine, which are involved in carbon recycling from photorespiration, and N-rich amino acids were predominantly increased in mETC mutants compared to the wild type. Taking together the physiological phenotypes of all examined mutants, our results suggest a connection between the limitation in the re-oxidation of reducing equivalents in the mitochondrial matrix and the induction of nitrate assimilation into free amino acids in the cytosol, which seems to be engaged as an additional sink for reducing power. The sdh2-1 mutant was less susceptible to C. higginsianum and did not show hampered salicylic acid (SA) accumulation as previously reported for SDH1 knock-down plants. The ROS burst remained unaffected in sdh2-1, emonstrating that subunit SDH2 is not involved in the control of ROS production and SA signaling by complex II. Moreover, the ndufs4 mutant showed only 20% of C. higginsianum colonization compared to wild type, with the ROS burst and the production of callose papillae being significantly increased compared to wild type. This indicates that a restriction of respiratory metabolism can positively affect pre-penetration resistance of Arabidopsis. Taking metabolite profiling data from all investigated mETC mutants, a strong positive correlation of resistance toward C. higginsianum with NADPH pool size, pyruvate contents, and other metabolites associated with redox poise and energy charge was evident, which fosters the hypothesis that limitations in the mETC can support resistance at post-penetration stages by improving the availability of metabolic power.