CHCHD4 binding affects the active site of apoptosis inducing factor (AIF): Structural determinants for allosteric regulation.

Apoptosis-inducing factor (AIF), which is confined to mitochondria of normal healthy cells, is the first identified caspase-independent cell death effector. Moreover, AIF is required for the optimal functioning of the respiratory chain machinery. Recent findings have revealed that AIF fulfills its pro-survival function by interacting with CHCHD4, a soluble mitochondrial protein which promotes the entrance and the oxidative folding of different proteins in the inner membrane space. Here, we report the crystal structure of the ternary complex involving the N-terminal 27-mer peptide of CHCHD4, NAD+, and AIF harboring its FAD (flavin adenine dinucleotide) prosthetic group in oxidized form. Combining this information with biophysical and biochemical data on the CHCHD4/AIF complex, we provide a detailed structural description of the interaction between the two proteins, validated by both chemical cross-linking mass spectrometry analysis and site-directed mutagenesis.

Response to microplastic exposure: An exploration into the sea urchin immune cell proteome.

It is now known that the Mediterranean Sea currently is one of the major hotspot for microplastics (MPs; < 5 mm) pollution and that the risks will be even more pronounced in the coming years. Thus, the in-depth study of the mechanisms underlying the MPs toxicity in key Mediterranean organisms, subjected to high anthropic pressures, has become a categorical imperative to pursue. Here, we explore for the first time the sea urchins immune cells profile combined to their proteome upon in vivo exposure (72 h) to different concentrations of polystyrene-microbeads (micro-PS) starting from relevant environmental concentrations (10, 50, 103, 104 MP/L). Every 24 h, immunological parameters were monitored. After 72 h, the abundance of MPs was examined in various organs and coelomocytes were collected for proteomic analysis based on a shotgun label free proteomic approach. While sea urchins treated with the lowest concentration tested (10 and 50 micro-PS/L) did not show the presence of micro-PS in any tissue, in the specimens exposed to the highest concentration (103 and 104 micro-PS) there was an internalisation of 9.75 ± 2.75 and 113.75 ± 34.5 MP/g, respectively. Proteomic analyses revealed that MPs exposure altered coelomocytes protein profile not only compared to the control group but also among the different micro-PS concentrations and these variations are micro-PS concentration dependent. The proteins exclusively expressed in the coelomocytes of specimens exposed to MPs are mainly metabolite interconversion enzymes, involved in cellular processes, indicating a severe alteration of the cellular metabolic pathways. Overall, these findings provide new insights on the mode of action of MPs in the sea urchin immune cells both at the molecular and cellular level.

The Neuroprotective Role of the GM1 Oligosaccharide, II3Neu5Ac-Gg4, in Neuroblastoma Cells.

Recently, we demonstrated that the GM1 oligosaccharide, II3Neu5Ac-Gg4 (OligoGM1), administered to cultured murine Neuro2a neuroblastoma cells interacts with the NGF receptor TrkA, leading to the activation of the ERK1/2 downstream pathway and to cell differentiation. To understand how the activation of the TrkA pathway is able to trigger key biochemical signaling, we performed a proteomic analysis on Neuro2a cells treated with 50 µM OligoGM1 for 24 h. Over 3000 proteins were identified. Among these, 324 proteins were exclusively expressed in OligoGM1-treated cells. Interestingly, several proteins expressed only in OligoGM1-treated cells are involved in biochemical mechanisms with a neuroprotective potential, reflecting the GM1 neuroprotective effect. In addition, we found that the exogenous administration of OligoGM1 reduced the cellular oxidative stress in Neuro2a cells and conferred protection against MPTP neurotoxicity. These results confirm and reinforce the idea that the molecular mechanisms underlying the GM1 neurotrophic and neuroprotective effects depend on its oligosaccharide chain, suggesting the activation of a positive signaling starting at plasma membrane level.