Senile plaques are a pathological hallmark of Alzheimer's disease (AD), yet the mechanism underlying their generation remains unknown. Beta-amyloid peptide (Aβ) is a major component of senile plaques. We analysed AD brain tissues with histochemistry, immunohistochemistry and fluorescence imaging to examine the neural, vascular or blood Aβ contribution to senile plaque development. We found little neural marker co-expression with plaque Aβ, while co-expression of blood markers, such as Haemin and ApoE, was abundant. The plaque cores were structured with vascular and glial proteins outside and blood metabolites inside, co-localizing with a characteristic of Hoechst staining-independent blue autofluorescence. Erythrocyte-interacting Aβ is linked to coagulation, elevated calcium and blue autofluorescence, and it is associated with intravascular haemolysis, atherosclerosis, cerebral amyloid angiopathy, microaneurysm, and often with Cathepsin D co-expression. We identified microaneurysms as major sites of amyloid formation. Our data suggest that senile plaques arise from Aβ- and Cathepsin D-enriched mixtures leaking out during intravascular haemolysis and microaneurysm rupture.

Small GTPases play essential roles in the organization of eukaryotic cells. In recent years, it has become clear that their intracellular functions result from intricate biochemical networks of the GTPase and their regulators that dynamically bind to a membrane surface. Due to the inherent complexities of their interactions, however, revealing the underlying mechanisms of action is often difficult to achieve from in vivo studies. This review summarizes in vitro reconstitution approaches developed to obtain a better mechanistic understanding of how small GTPase activities are regulated in space and time.

Prolonged metabolic stress can lead to severe pathologies. In metabolically challenged primary fibroblasts, we assigned a novel role for the poorly characterized miR-4734 in restricting ATF4 and IRE1-mediated upregulation of a set of proinflammatory cytokines and endoplasmic reticulum stress-associated genes. Conversely, inhibition of this miRNA augmented the expression of those genes. Mechanistically, miR-4734 was found to restrict the expression of the transcriptional activator NF-kappa-B inhibitor zeta (NFKBIZ), which is required for optimal expression of the proinflammatory genes and whose mRNA is targeted directly by miR-4734. Concordantly, overexpression of NFKBIZ compromised the effects of miR-4734, underscoring the importance of this direct targeting. As the effects of miR-4734 were evident under stress but not under basal conditions, it may possess therapeutic utility towards alleviating stress-induced pathologies.

Paraclostridial mosquitocidal protein 1 (PMP1) is a member of the clostridial neurotoxin (CNT) family, which includes botulinum and tetanus neurotoxins. PMP1 has unique selectivity for anopheline mosquitos and is the only known member of the family that targets insects. PMP1 is encoded in an orfX gene cluster, which in addition to the toxin, consists of OrfX1, OrfX2, OrfX3, P47 and NTNH, which have been shown to aid in PMP1 toxicity. We here show that OrfX1 and OrfX3 form a complex and present its structure at 2.7 Å. The OrfX1-OrfX3 complex mimics the structure of full-length OrfX2 and belongs to the lipid-binding TULIP protein superfamily. With this report, the structures of all proteins encoded in the orfX gene cluster of CNTs are now determined.

Small GTPases act as molecular switches and control numerous cellular processes by their binding and hydrolysis of guanosine triphosphate (GTP). The activity of small GTPases is coordinated by guanine nucleotide exchange factors (GEFs) and GTPase activating proteins (GAPs). Recent structural and functional studies have characterized a subset of GAPs whose catalytic units consist of longin domains. Longin domain containing GAPs regulate small GTPases that facilitate nutrient signalling, autophagy, vesicular trafficking and lysosome homeostasis. All known examples in this GAP family function as part of larger multiprotein complexes. The three characterized mammalian protein complexes in this class are FLCN:FNIP, GATOR1 and C9orf72:SMCR8. Each complex carries out a unique cellular function by regulating distinct small GTPases. In this article, we explore the roles of longin domain GAPs in nutrient sensing, membrane dynamic, vesicular trafficking and disease. Through a structural lens, we examine the mechanism of each longin domain GAP and highlight potential therapeutic applications.