Reconfiguring k-path vertex covers

A vertex subset $I$ of a graph $G$ is called a $k$-path vertex cover if every path on $k$ vertices in $G$ contains at least one vertex from $I$. The \textsc{$k$-Path Vertex Cover Reconfiguration ($k$-PVCR)} problem asks if one can transform one $k$-path vertex cover into another via a sequence of $k$-path vertex covers where each intermediate member is obtained from its predecessor by applying a given reconfiguration rule exactly once. We investigate the computational complexity of \textsc{$k$-PVCR} from the viewpoint of graph classes under the well-known reconfiguration rules: $\mathsf{TS}$, $\mathsf{TJ}$, and $\mathsf{TAR}$. The problem for $k=2$, known as the \textsc{Vertex Cover Reconfiguration (VCR)} problem, has been well-studied in the literature. We show that certain known hardness results for \textsc{VCR} on different graph classes including planar graphs, bounded bandwidth graphs, chordal graphs, and bipartite graphs, can be extended for \textsc{$k$-PVCR}. In particular, we prove a complexity dichotomy for \textsc{$k$-PVCR} on general graphs: on those whose maximum degree is $3$ (and even planar), the problem is $\mathtt{PSPACE}$-complete, while on those whose maximum degree is $2$ (i.e., paths and cycles), the problem can be solved in polynomial time. Additionally, we also design polynomial-time algorithms for \textsc{$k$-PVCR} on trees under each of $\mathsf{TJ}$ and $\mathsf{TAR}$. Moreover, on paths, cycles, and trees, we describe how one can construct a reconfiguration sequence between two given $k$-path vertex covers in a yes-instance. In particular, on paths, our constructed reconfiguration sequence is shortest.Comment: 29 pages, 4 figures, to appear in WALCOM 202

Reconfiguring k-Path Vertex Covers

A vertex subset I of a graph G is called a k-path vertex cover if every path on k vertices in G contains at least one vertex from I. The K-PATH VERTEX COVER RECONFIGURATION (K-PVCR) problem asks if one can transform one k-path vertex cover into another via a sequence of k-path vertex covers where each intermediate member is obtained from its predecessor by applying a given reconfiguration rule exactly once. We investigate the computational complexity of K-PVCR from the viewpoint of graph classes under the well-known reconfiguration rules: TS, TJ, and TAR. The problem for k=2, known as the VERTEX COVER RECONFIGURATION (VCR) problem, has been well-studied in the literature. We show that certain known hardness results for VCR on different graph classes can be extended for K-PVCR. In particular, we prove a complexity dichotomy for K-PVCR on general graphs: on those whose maximum degree is three (and even planar), the problem is PSPACE-complete, while on those whose maximum degree is two (i.e., paths and cycles), the problem can be solved in polynomial time. Additionally, we also design polynomial-time algorithms for K-PVCR on trees under each of TJ and TAR. Moreover, on paths, cycles, and trees, we describe how one can construct a reconfiguration sequence between two given k-path vertex covers in a yes-instance. In particular, on paths, our constructed reconfiguration sequence is shortest

Cellular Inhibitor of Apoptosis (cIAP)-mediated ubiquitination of Phosphofurin Acidic Cluster Sorting protein 2 (PACS-2) negatively regulates Tumor necrosis factor-Related Apoptosis-Inducing Ligand (TRAIL) cytotoxicity

Lysosomal membrane permeabilization is an essential step in TRAIL-induced apoptosis of liver cancer cell lines. TRAIL-induced lysosomal membrane permeabilization is mediated by the multifunctional sorting protein PACS-2 and repressed by the E3 ligases cIAP-1 and cIAP-2. Despite the opposing roles for PACS-2 and cIAPs in TRAIL-induced apoptosis, an interaction between these proteins has yet to be examined. Herein, we report that cIAP-1 and cIAP-2 confer TRAIL resistance to hepatobiliary cancer cell lines by reducing PACS-2 levels. Under basal conditions, PACS-2 underwent K48-linked polyubiquitination, resulting in PACS-2 proteasomal degradation. Biochemical assays showed cIAP-1 and cIAP-2 interacted with PACS-2 in vitro and co-immunoprecipitation studies demonstrated that the two cIAPs bound PACS-2 in vivo. More importantly, both cIAP-1 and cIAP-2 directly mediated PACS-2 ubiquitination in a cell-free assay. Single c-Iap-1 or c-Iap-2 gene knock-outs in mouse hepatocytes did not lead to PACS-2 accumulation. However, deletion of both cIAP-1 and cIAP-2 reduced PACS-2 ubiquitination, which increased PACS-2 levels and sensitized HuH-7 cells to TRAIL-induced lysosomal membrane permeabilization and apoptosis. Correspondingly, deletion of cIAPs sensitized wild-type, but not PACS-2-deficient hepatocarcinoma cells or Pacs-2-/- mouse hepatocytes to TRAIL-induced apoptosis. Together, these data suggest cIAPs constitutively downregulate PACS-2 by polyubiquitination and proteasomal degradation, thereby restraining TRAIL-induced killing of liver cancer cells. © 2014 Guicciardi et al

Characterization and application of two RANK-specific antibodies with different biological activities.

Antibodies play an important role in therapy and investigative biomedical research. The TNF-family member Receptor Activator of NF-κB (RANK) is known for its role in bone homeostasis and is increasingly recognized as a central player in immune regulation and epithelial cell activation. However, the study of RANK biology has been hampered by missing or insufficient characterization of high affinity tools that recognize RANK. Here, we present a careful description and comparison of two antibodies, RANK-02 obtained by phage display (Newa, 2014 [1]) and R12-31 generated by immunization (Kamijo, 2006 [2]). We found that both antibodies recognized mouse RANK with high affinity, while RANK-02 and R12-31 recognized human RANK with high and lower affinities, respectively. Using a cell apoptosis assay based on stimulation of a RANK:Fas fusion protein, and a cellular NF-κB signaling assay, we showed that R12-31 was agonist for both species. R12-31 interfered little or not at all with the binding of RANKL to RANK, in contrast to RANK-02 that efficiently prevented this interaction. Depending on the assay and species, RANK-02 was either a weak agonist or a partial antagonist of RANK. Both antibodies recognized human Langerhans cells, previously shown to express RANK, while dermal dendritic cells were poorly labeled. In vivo R12-31 agonist activity was demonstrated by its ability to induce the formation of intestinal villous microfold cells in mice. This characterization of two monoclonal antibodies should now allow better evaluation of their application as therapeutic reagents and investigative tools

The architecture of EMC reveals a path for membrane protein insertion.

Funder: Boehringer Ingelheim Fonds; FundRef: http://dx.doi.org/10.13039/501100001645Funder: Naito Foundation; FundRef: http://dx.doi.org/10.13039/100007428Funder: Japanese Biochemical SocietyApproximately 25% of eukaryotic genes code for integral membrane proteins that are assembled at the endoplasmic reticulum. An abundant and widely conserved multi-protein complex termed EMC has been implicated in membrane protein biogenesis, but its mechanism of action is poorly understood. Here, we define the composition and architecture of human EMC using biochemical assays, crystallography of individual subunits, site-specific photocrosslinking, and cryo-EM reconstruction. Our results suggest that EMC's cytosolic domain contains a large, moderately hydrophobic vestibule that can bind a substrate's transmembrane domain (TMD). The cytosolic vestibule leads into a lumenally-sealed, lipid-exposed intramembrane groove large enough to accommodate a single substrate TMD. A gap between the cytosolic vestibule and intramembrane groove provides a potential path for substrate egress from EMC. These findings suggest how EMC facilitates energy-independent membrane insertion of TMDs, explain why only short lumenal domains are translocated by EMC, and constrain models of EMC's proposed chaperone function

The architecture of EMC reveals a path for membrane protein insertion

Approximately 25% of eukaryotic genes code for integral membrane proteins that are assembled at the endoplasmic reticulum. An abundant and widely conserved multi-protein complex termed EMC has been implicated in membrane protein biogenesis, but its mechanism of action is poorly understood. Here, we define the composition and architecture of human EMC using biochemical assays, crystallography of individual subunits, site-specific photocrosslinking, and cryo-EM reconstruction. Our results suggest that EMC’s cytosolic domain contains a large, moderately hydrophobic vestibule that can bind a substrate’s transmembrane domain (TMD). The cytosolic vestibule leads into a lumenally-sealed, lipid-exposed intramembrane groove large enough to accommodate a single substrate TMD. A gap between the cytosolic vestibule and intramembrane groove provides a potential path for substrate egress from EMC. These findings suggest how EMC facilitates energy-independent membrane insertion of TMDs, explain why only short lumenal domains are translocated by EMC, and constrain models of EMC’s proposed chaperone function

The architecture of EMC reveals a path for membrane protein insertion

Approximately 25% of eukaryotic genes code for integral membrane proteins that are assembled at the endoplasmic reticulum. An abundant and widely conserved multi-protein complex termed EMC has been implicated in membrane protein biogenesis, but its mechanism of action is poorly understood. Here, we define the composition and architecture of human EMC using biochemical assays, crystallography of individual subunits, site-specific photocrosslinking, and cryo-EM reconstruction. Our results suggest that EMC’s cytosolic domain contains a large, moderately hydrophobic vestibule that can bind a substrate’s transmembrane domain (TMD). The cytosolic vestibule leads into a lumenally-sealed, lipid-exposed intramembrane groove large enough to accommodate a single substrate TMD. A gap between the cytosolic vestibule and intramembrane groove provides a potential path for substrate egress from EMC. These findings suggest how EMC facilitates energy-independent membrane insertion of TMDs, explain why only short lumenal domains are translocated by EMC, and constrain models of EMC’s proposed chaperone function

The architecture of EMC reveals a path for membrane protein insertion

Approximately 25% of eukaryotic genes code for integral membrane proteins that are assembled at the endoplasmic reticulum. An abundant and widely conserved multi-protein complex termed EMC has been implicated in membrane protein biogenesis, but its mechanism of action is poorly understood. Here, we define the composition and architecture of human EMC using biochemical assays, crystallography of individual subunits, site-specific photocrosslinking, and cryo-EM reconstruction. Our results suggest that EMC’s cytosolic domain contains a large, moderately hydrophobic vestibule that can bind a substrate’s transmembrane domain (TMD). The cytosolic vestibule leads into a lumenally-sealed, lipid-exposed intramembrane groove large enough to accommodate a single substrate TMD. A gap between the cytosolic vestibule and intramembrane groove provides a potential path for substrate egress from EMC. These findings suggest how EMC facilitates energy-independent membrane insertion of TMDs, explain why only short lumenal domains are translocated by EMC, and constrain models of EMC’s proposed chaperone function

Dimerization and nuclear entry of mPER proteins in mammalian cells

Nuclear entry of circadian oscillatory gene products is a key step for the generation of a 24-hr cycle of the biological clock. We have examined nuclear import of clock proteins of the mammalian period gene family and the effect of serum shock, which induces a synchronous clock in cultured cells. Previously, mCRY1 and mCRY2 have been found to complex with PER proteins leading to nuclear import. Here we report that nuclear translocation of mPER1 and mPER2 (1) involves physical interactions with mPER3, (2) is accelerated by serum treatment, and (3) still occurs in mCry1/mCry2 double-deficient cells lacking a functional biological clock. Moreover, nuclear localization of endogenous mPER1 was observed in cultured mCry1/mCry2 double-deficient cells as well as in the liver and the suprachiasmatic nuclei (SCN) of mCry1/mCry2 double-mutant mice. This indicates that nuclear translocation of at least mPER1 also can occur under physiological conditions (i.e., in the intact mouse) in the absence of any CRY protein. The mPER3 amino acid sequence predicts the presence of a cytoplasmic localization domain (CLD) and a nuclear localization signal (NLS). Deletion analysis suggests that the interplay of the CLD and NLS proposed to regulate nuclear entry of PER in Drosophila is conserved in mammals, but with the novel twist that mPER3 can act as the dimerizing partner