Novel human and mouse genes encoding a shank-interacting protein and its upregulation in gastric fundus ofW/WVmouse

Mixed-lineage kinase domain-like protein (MLKL) is essential for TNF-α–induced necroptosis. How MLKL promotes cell death is still under debate. Here we report that MLKL forms SDS-resistant, disulfide bond-dependent polymers during necroptosis in both human and mouse cells. MLKL polymers are independent of receptor-interacting protein kinase 1 and 3 (RIPK1/RIPK3) fibers. Large MLKL polymers are more than 2 million Da and are resistant to proteinase K digestion. MLKL polymers are fibers 5 nm in diameter under electron microscopy. Furthermore, the recombinant N-terminal domain of MLKL forms amyloid-like fibers and binds Congo red dye. MLKL mutants that cannot form polymers also fail to induce necroptosis efficiently. Finally, the compound necrosulfonamide conjugates cysteine 86 of human MLKL and blocks MLKL polymer formation and subsequent cell death. These results demonstrate that disulfide bond-dependent, amyloid-like MLKL polymers are necessary and sufficient to induce necroptosis.

Download Full-text

Identification of an alpha-tubulin mutant of fission yeast from gamma-tubulin-interacting protein screening: genetic evidence for alpha-/gamma-tubulin interaction

Journal of Cell Science ◽

10.1242/jcs.113.24.4557 ◽

2000 ◽

Vol 113 (24) ◽

pp. 4557-4562 ◽

Cited By ~ 1

Author(s):

A. Takeoka ◽

M. Shimizu ◽

T. Horio

Keyword(s):

Fission Yeast ◽

Central Element ◽

Direct Interaction ◽

Interacting Proteins ◽

Microtubule Nucleation ◽

Mutation Site ◽

Interacting Protein ◽

Alpha Tubulin ◽

Genes Encoding ◽

Gamma Tubulin

gamma-Tubulin has been determined to be a central element of microtubule nucleation and, thus, indispensable for cellular organization of the microtubule. Utilizing the fact that human gamma-tubulin can function in the fission yeast Schizosaccharomyces pombe, we have generated a unique mutant screening procedure which can specifically select mutants of genes encoding gamma-tubulin-interacting proteins. One of the isolated mutants, cs76, turned out to carry a mutation in the alpha 1-tubulin gene (nda2(+)). This result suggests a direct interaction between the alpha- and gamma-tubulins. We located the mutation site in the nda2 gene and characterized the mutant phenotype. Our results demonstrate the importance of the alpha-/gamma-tubulin interaction in microtubule nucleation and should complement previous knowledge.

Download Full-text

cDNA cloning, expression profile, and genomic structure of human and mouse RNF10/Rnf 10 genes, encoding a novel RING finger protein

Journal of Human Genetics ◽

10.1007/s100380050007 ◽

2000 ◽

Vol 45 (1) ◽

pp. 38-42 ◽

Cited By ~ 8

Author(s):

N. Seki ◽

A. Hattori ◽

S. Sugano ◽

M. Muramatsu ◽

T. Saito

Keyword(s):

Cdna Cloning ◽

Expression Profile ◽

Genomic Structure ◽

Ring Finger ◽

Ring Finger Protein ◽

Finger Protein ◽

Genes Encoding ◽

Human And Mouse

Download Full-text

Molecular and Functional Analyses of the Human and Mouse Genes Encoding AFG3L1, a Mitochondrial Metalloprotease Homologous to the Human Spastic Paraplegia Protein

Genomics ◽

10.1006/geno.2001.6560 ◽

2001 ◽

Vol 76 (1-3) ◽

pp. 58-65 ◽

Cited By ~ 34

Author(s):

Gabriel Kremmidiotis ◽

Alison E. Gardner ◽

Chatri Settasatian ◽

Anna Savoia ◽

Grant R. Sutherland ◽

...

Keyword(s):

Spastic Paraplegia ◽

Genes Encoding ◽

Functional Analyses ◽

Human And Mouse

Download Full-text

Diverse Sequence Determinants Control Human and Mouse Receptor Interacting Protein 3 (RIP3) and Mixed Lineage Kinase domain-Like (MLKL) Interaction in Necroptotic Signaling

Journal of Biological Chemistry ◽

10.1074/jbc.m112.435545 ◽

2013 ◽

Vol 288 (23) ◽

pp. 16247-16261 ◽

Cited By ~ 138

Author(s):

Wanze Chen ◽

Zhenru Zhou ◽

Lisheng Li ◽

Chuan-Qi Zhong ◽

Xinru Zheng ◽

...

Keyword(s):

Protein Kinase ◽

Kinase Domain ◽

Phosphorylation Sites ◽

Flanking Sequence ◽

Interacting Protein ◽

Mixed Lineage Kinase ◽

Signaling Complex ◽

Evolutionarily Conserved ◽

Mouse Cells ◽

Human And Mouse

Receptor interacting protein 3 (RIP3) is a protein kinase essential for TNF-induced necroptosis. Phosphorylation on Ser-227 in human RIP3 (hRIP3) is required for its interaction with human mixed lineage kinase domain-like (MLKL) in the necrosome, a signaling complex induced by TNF stimulation. RIP1 and RIP3 mediate necrosome aggregation leading to the formation of amyloid-like signaling complexes. We found that TNF induces Thr-231 and Ser-232 phosphorylation in mouse RIP3 (mRIP3) and this phosphorylation is required for mRIP3 to interact with mMLKL. Ser-232 in mRIP3 corresponds to Ser-227 in hRIP3, whereas Thr-231 is not conserved in hRIP3. Although the RIP3-MLKL interaction is required for necroptosis in both human and mouse cells, hRIP3 does not interact with mMLKL and mRIP3 cannot bind to hMLKL. The species specificity of the RIP3-MLKL interaction is primarily determined by the sequence differences in the phosphorylation sites and the flanking sequence around the phosphorylation sites in hRIP3 and mRIP3. It appears that the RIP3-MLKL interaction has been selected as an evolutionarily conserved mechanism in mediating necroptosis signaling despite that differing structural and mechanistic bases for this interaction emerged simultaneously in different organisms. In addition, we further revealed that the interaction of RIP3 with MLKL prevented massive abnormal RIP3 aggregation, and therefore should be crucial for formation of the amyloid signaling complex of necrosomes. We also found that the interaction between RIP3 and MLKL is required for the translocation of necrosomes to mitochondria-associated membranes. Our data demonstrate the importance of the RIP3-MLKL interaction in the formation of functional necrosomes and suggest that translocation of necrosomes to mitochondria-associated membranes is essential for necroptosis signaling.

Download Full-text

Aldose Reductases Influence Prostaglandin F2α Levels and Adipocyte Differentiation in Male Mouse and Human Species

Endocrinology ◽

10.1210/en.2014-1750 ◽

2015 ◽

Vol 156 (5) ◽

pp. 1671-1684 ◽

Cited By ~ 5

Author(s):

Emilie Pastel ◽

Jean-Christophe Pointud ◽

Gaëlle Loubeau ◽

Christian Dani ◽

Karem Slim ◽

...

Keyword(s):

Aldose Reductase ◽

Adipocyte Differentiation ◽

Stromal Vascular Fraction ◽

Prostaglandin F2α ◽

Early Differentiation ◽

Reductase Inhibitors ◽

Expression Of Genes ◽

Gene Ablation ◽

Genes Encoding ◽

Human And Mouse

Aldose reductases (AKR1B) are widely expressed oxidoreductases whose physiological function remains elusive. Some isoforms are genuine prostaglandin F2α (PGF2α) synthases, suggesting they might influence adipose homeostasis because PGF2α inhibits adipogenesis. This was shown by Akr1b7 gene ablation in the mouse, which resulted in increased adiposity related to a lower PGF2α content in fat. Yet humans have no ortholog gene for Akr1b7, so the role of aldose reductases in human adipose homeostasis remains to be explored. We analyzed expression of genes encoding human and mouse aldose reductase isoforms in adipose tissues and differentiating adipocytes to assess conserved mechanisms regulating PGF2α synthesis and adipogenesis. The Akr1b3 gene encoded the most abundant isoform in mouse adipose tissue, whereas Akr1b7 encoded the only isoform enriched in the stromal vascular fraction. Most mouse aldose reductase gene expression peaked in early adipogenesis of 3T3-L1 cells and diminished with differentiation. In contrast with its mouse ortholog Akr1b3, AKR1B1 expression increased throughout differentiation of human multipotent adipose-derived stem cells, paralleling PGF2α release, whereas PGF2α receptor (FP) levels collapsed in early differentiation. Pharmacological inhibition of aldose reductase using Statil altered PGF2α production and enhanced human multipotent adipose-derived stem adipocyte differentiation. As expected, the adipogenic effects of Statil were counteracted by an FP agonist (cloprostenol). Thus, in both species aldose reductase-dependent PGF2α production could be important in early differentiation to restrict adipogenesis. PGF2α antiadipogenic signaling could then be toned down through the FP receptor or aldose reductases down-regulation in human and mouse cells, respectively. Our data suggest that aldose reductase inhibitors could have obesogenic potential.

Download Full-text

The Amyloid Structure of Mouse RIPK3 (Receptor Interacting Protein Kinase 3) in Cell Necroptosis

10.1101/2020.07.10.196980 ◽

2020 ◽

Author(s):

Xia-lian Wu ◽

Hong Hu ◽

Xing-qi Dong ◽

Jing Zhang ◽

Jian Wang ◽

...

Keyword(s):

Structural Transformation ◽

Self Assembly ◽

Structural Integrity ◽

Immune Defense ◽

Dynamics Simulation ◽

Amyloid Formation ◽

Interacting Protein ◽

Functional Studies ◽

First Time ◽

Human And Mouse

ABSTRACTRIPK3 amyloid complex plays crucial roles in execution of TNF-induced necroptosis and in response to immune defense in both human and mouse. We have structurally characterized the mouse RIPK3 homogeneous self-assembly using solid-state NMR, illustrating a well-ordered N-shaped amyloid core structure featured with 3 parallel in-register β-sheets. The structure is different from previously published human RIPK1/RIPK3 hetero-amyloid complex. Functional studies indicate both RIPK1-RIPK3 binding and RIPK3 amyloid formation are essential but not sufficient for RIPK3-mediated necroptosis. The structural integrity of RIPK3 fibril with three β-strands is necessary for the signaling. Molecular dynamics simulation of the mouse RIPK1/RIPK3 model indicates less stable for the hetero-amyloid to adopt RIPK3 fibril conformation, suggesting a structural transformation of RIPK3 from RIPK1-RIPK3 binding to RIPK3 amyloid formation. This structural transformation is revealed for the first time, providing a missing link connecting RIPK1-RIPK3 binding to RIPK3 homo-oligomer formation in the signal transduction.

Download Full-text

Development of an RNA interference (RNAi) gene knockdown protocol in the anaerobic gut fungus Pecoramyces ruminantium strain C1A

10.7287/peerj.preprints.3177 ◽

2017 ◽

Author(s):

Shelby S Calkins ◽

Nicole C Elledge ◽

Stephen M. Marek ◽

M B. Couger ◽

Mostafa S Elshahed ◽

...

Keyword(s):

Rna Interference ◽

Gene Silencing ◽

Neurospora Crassa ◽

Dna Helicase ◽

Small Interfering Rnas ◽

Gene Encoding ◽

Interacting Protein ◽

Genes Encoding ◽

Lactate Levels ◽

Electron Sink

Members of the anaerobic gut fungi (AGF) reside in rumen, hindgut, and feces of ruminant and non-ruminant herbivorous mammals and reptilian herbivores. No protocols for gene insertion, deletion, silencing, or mutation are currently available for the AGF, rendering gene-targeted molecular biological manipulations unfeasible. Here, we developed and optimized an RNA interference (RNAi)-based protocol for targeted gene silencing in the anaerobic gut fungus Pecoramyces ruminantium strain C1A. Analysis of the C1A genome identified genes encoding enzymes required for RNA silencing in fungi (Dicer, Argonaute, Neurospora crassa QDE-3 homolog DNA helicase, Argonaute-interacting protein, and Neurospora crassa QIP homolog exonuclease); and the competency of C1A germinating spores for RNA uptake was confirmed using fluorescently labeled small interfering RNAs (siRNA). Addition of chemically-synthesized siRNAs targeting D-lactate dehydrogenase (ldhD) gene to C1A germinating spores resulted in marked target gene silencing; as evident by significantly lower ldhD transcriptional levels, a marked reduction in the D-LDH specific enzymatic activity in intracellular protein extracts, and a reduction in D-lactate levels accumulating in the culture supernatant. Comparative transcriptomic analysis of untreated versus siRNA-treated cultures identified a few off-target siRNA-mediated gene silencing effects. As well, significant differential up-regulation of the gene encoding NAD-dependent 2-hydroxyacid dehydrogenase (Pfam00389) in siRNA-treated C1A cultures was observed, which could possibly compensate for loss of D-LDH as an electron sink mechanism in C1A. The results demonstrate the feasibility of RNAi in anaerobic fungi, and opens the door for gene silencing-based studies in this fungal clade.

Download Full-text