NETosis, the process wherein neutrophils release highly decondensed chromatin called neutrophil extracellular traps (NETs), has gained much attention as an alternative means of killing bacteria. In vivo, NETs are induced by bacteria and pro-inflammatory cytokines. We have reported that peptidylarginine deiminase 4 (PAD4), an enzyme that converts Arg or monomethyl-Arg to citrulline in histones, is essential for NET formation. The areas of extensive chromatin decondensation along the NETs were rich in histone citrullination. Here, upon investigating the effect of global citrullination in cultured cells, we discovered that PAD4 overexpression in osteosarcoma U2OS cells induces extensive chromatin decondensation independent of apoptosis. The highly decondensed chromatin is released to the extracellular space and stained strongly by a histone citrulline-specific antibody. The structure of the decondensed chromatin is reminiscent of NETs but is unique in that it occurs without stimulation of cells with pro-inflammatory cytokines and bacteria. Furthermore, histone citrullination during chromatin decondensation can dissociate heterochromatin protein 1 beta (HP1β) thereby offering a new molecular mechanism for understanding how citrullination regulates chromatin function. Taken together, our study suggests that PAD4 mediated citrullination induces chromatin decondensation, implicating its essential role in NET formation under physiological conditions in neutrophils.

Summary We have recently demonstrated that reactive glial cells can be directly reprogrammed into functional neurons by single neural transcription factor NeuroD1. Here we report that a combination of small molecules can also reprogram human astrocytes in culture into fully functional neurons. We demonstrate that sequential exposure of human astrocytes to a cocktail of 9 small molecules that inhibit glial but activate neuronal signaling pathways can successfully reprogram astrocytes into neurons in 8-10 days. This chemical reprogramming is mediated through epigenetic regulation and involves transcriptional activation of NEUROD1 and NEUROGENIN2. The human astrocyte-converted neurons can survive for >5 months in culture and form functional synaptic networks with synchronous burst activities. The chemical reprogrammed human neurons can also survive for >1 month in the mouse brain in vivo and integrate into local circuits. Our study opens a new avenue using chemical compounds to reprogram reactive glial cells into functional neurons.

Background: Histone citrullination by PAD4 regulates tumor suppressor gene expression. Results:The novel PAD inhibitor YW3-56 inhibits cancerous growth by perturbing autophagy and regulating the SESN2-mTORC1 signaling axis. Conclusion: YW3-56 regulates the SESN2-mTORC1 autophagy pathway as one of its anticancer mechanisms. Significance: This study identifies a novel function of PAD4 in the autophagy pathway and developed potent PAD inhibitors for future cancer research.

Background: Cofilin rods are associated with Alzheimer disease, but their pathological significance is unclear. Results: Time-lapse imaging revealed that cofilin rods inhibit the movement of mitochondria and early endosomes. Cofilin rods reduce dendritic spines and impair synaptic transmission. Cofilin rods are discovered in senile rat brains. Conclusion: Cofilin rods block intracellular transport and induce synaptic loss. Significance: Our work identifies a signaling pathway underlying neurodegeneration and brain aging.