A dual-receptor T-cell platform with Ab-TCR and costimulatory receptor achieves specificity and potency against AML.

ABSTRACT: Chimeric antigen receptor T-cell (CAR T) therapy has produced remarkable clinical responses in B-cell neoplasms. However, many challenges limit this class of agents for the treatment of other cancer types, in particular the lack of tumor-selective antigens for solid tumors and other hematological malignancies, such as acute myeloid leukemia (AML), which may be addressed without significant risk of severe toxicities while providing sufficient abundance for efficient tumor suppression. One approach to overcome this hurdle is dual targeting by an antibody-T-cell receptor (AbTCR) and a chimeric costimulatory signaling receptor (CSR) to 2 different antigens, in which both antigens are found together on the cancer cells but not together on normal cells. To explore this proof of concept in AML, we engineered a new T-cell format targeting Wilms tumor 1 protein (WT1) and CD33; both are highly expressed on most AML cells. Using an AbTCR comprising a newly developed TCR-mimic monoclonal antibody against the WT1 RMFPNAPYL (RMF) epitope/HLA-A2 complex, ESK2, and a secondary CSR comprising a single-chain variable fragment directed to CD33 linked to a truncated CD28 costimulatory fragment, this unique platform confers specific T-cell cytotoxicity to the AML cells while sparing healthy hematopoietic cells, including CD33+ myelomonocytic normal cells. These data suggest that this new platform, named AbTCR-CSR, through the combination of a AbTCR CAR and CSR could be an effective strategy to reduce toxicity and improve specificity and clinical outcomes in adoptive T-cell therapy in AML.

MyD88 is required for satellite cell-mediated myofiber regeneration in dystrophin-deficient mdx mice.

Duchenne muscular dystrophy (DMD), caused by mutations in the dystrophin gene, leads to severe muscle wasting and eventual death of the afflicted individuals, primarily due to respiratory failure. Deficit in myofiber regeneration, potentially due to an exhaustion of satellite cells, is one of the major pathological features of DMD. Myeloid differentiation primary response 88 (MyD88) is an adaptor protein that mediates activation of various inflammatory pathways in response to signaling from Toll-like receptors and interleukin-1 receptor. MyD88 also regulates cellular survival, proliferation and differentiation in a cell-autonomous manner. However, the role of MyD88 in satellite stem cell homeostasis and function in dystrophic muscle remains unknown. In this study, we demonstrate that tamoxifen-inducible deletion of MyD88 in satellite cells causes loss of skeletal muscle mass and strength in the mdx mouse model of DMD. Satellite cell-specific deletion of MyD88 inhibits myofiber regeneration and stimulates fibrogenesis in dystrophic muscle of mdx mice. Deletion of MyD88 also reduces the number of satellite cells and inhibits their fusion with injured myofibers in dystrophic muscle of mdx mice. Ablation of MyD88 in satellite cells increases the markers of M2 macrophages without having any significant effect on M1 macrophages and expression of inflammatory cytokines. Finally, we found that satellite cell-specific deletion of MyD88 leads to aberrant activation of Notch and Wnt signaling in skeletal muscle of mdx mice. Collectively, our results demonstrate that MyD88-mediated signaling in satellite cells is essential for the regeneration of injured myofibers in dystrophic muscle of mdx mice.

PERK regulates skeletal muscle mass and contractile function in adult mice.

Skeletal muscle mass is regulated by the coordinated activation of several anabolic and catabolic pathways. The endoplasmic reticulum (ER) is a major site of protein folding and a reservoir for calcium ions. Accretion of misfolded proteins or depletion in calcium concentration causes stress in the ER, which leads to the activation of a signaling network known as the unfolded protein response (UPR). In the present study, we investigated the role of the protein kinase R-like endoplasmic reticulum kinase (PERK) arm of the UPR in the regulation of skeletal muscle mass and function in naive conditions and in a mouse model of cancer cachexia. Our results demonstrate that the targeted inducible deletion of PERK reduces skeletal muscle mass, strength, and force production during isometric contractions. Deletion of PERK also causes a slow-to-fast fiber type transition in skeletal muscle. Furthermore, short hairpin RNA-mediated knockdown or pharmacologic inhibition of PERK leads to atrophy in cultured myotubes. While increasing the rate of protein synthesis, the targeted deletion of PERK leads to the increased expression of components of the ubiquitin-proteasome system and autophagy in skeletal muscle. Ablation of PERK also increases the activation of calpains and deregulates the gene expression of the members of the FGF19 subfamily. Furthermore, the targeted deletion of PERK increases muscle wasting in Lewis lung carcinoma tumor-bearing mice. Our findings suggest that the PERK arm of the UPR is essential for the maintenance of skeletal muscle mass and function in adult mice.-Gallot, Y. S., Bohnert, K. R., Straughn, A. R., Xiong, G., Hindi, S. M., Kumar, A. PERK regulates skeletal muscle mass and contractile function in adult mice.

Pectin Biosynthesis Is Critical for Cell Wall Integrity and Immunity in Arabidopsis thaliana.

Plant cell walls are important barriers against microbial pathogens. Cell walls of Arabidopsis thaliana leaves contain three major types of polysaccharides: cellulose, various hemicelluloses, and pectins. UDP-D-galacturonic acid, the key building block of pectins, is produced from the precursor UDP-D-glucuronic acid by the action of glucuronate 4-epimerases (GAEs). Pseudomonas syringae pv maculicola ES4326 (Pma ES4326) repressed expression of GAE1 and GAE6 in Arabidopsis, and immunity to Pma ES4326 was compromised in gae6 and gae1 gae6 mutant plants. These plants had brittle leaves and cell walls of leaves had less galacturonic acid. Resistance to specific Botrytis cinerea isolates was also compromised in gae1 gae6 double mutant plants. Although oligogalacturonide (OG)-induced immune signaling was unaltered in gae1 gae6 mutant plants, immune signaling induced by a commercial pectinase, macerozyme, was reduced. Macerozyme treatment or infection with B. cinerea released less soluble uronic acid, likely reflecting fewer OGs, from gae1 gae6 cell walls than from wild-type Col-0. Although both OGs and macerozyme-induced immunity to B. cinerea in Col-0, only OGs also induced immunity in gae1 gae6. Pectin is thus an important contributor to plant immunity, and this is due at least in part to the induction of immune responses by soluble pectin, likely OGs, that are released during plant-pathogen interactions.

Identification and Characterization of a Golgi-Localized UDP-Xylose Transporter Family from Arabidopsis.

Most glycosylation reactions require activated glycosyl donors in the form of nucleotide sugars to drive processes such as posttranslational modifications and polysaccharide biosynthesis. Most plant cell wall polysaccharides are biosynthesized in the Golgi apparatus from cytosolic-derived nucleotide sugars, which are actively transferred into the Golgi lumen by nucleotide sugar transporters (NSTs). An exception is UDP-xylose, which is biosynthesized in both the cytosol and the Golgi lumen by a family of UDP-xylose synthases. The NST-based transport of UDP-xylose into the Golgi lumen would appear to be redundant. However, employing a recently developed approach, we identified three UDP-xylose transporters in the Arabidopsis thaliana NST family and designated them UDP-XYLOSE TRANSPORTER1 (UXT1) to UXT3. All three transporters localize to the Golgi apparatus, and UXT1 also localizes to the endoplasmic reticulum. Mutants in UXT1 exhibit ∼30% reduction in xylose in stem cell walls. These findings support the importance of the cytosolic UDP-xylose pool and UDP-xylose transporters in cell wall biosynthesis.

Inhibition of ER stress and unfolding protein response pathways causes skeletal muscle wasting during cancer cachexia.

Cachexia is a devastating syndrome that causes morbidity and mortality in a large number of patients with cancer. However, the mechanisms of cancer cachexia remain poorly understood. Accumulation of misfolded proteins in the endoplasmic reticulum (ER) causes stress. The ER responds to this stress through activating certain pathways commonly known as the unfolding protein response (UPR). The main function of UPR is to restore homeostasis, but excessive or prolonged activation of UPR can lead to pathologic conditions. In this study, we examined the role of ER stress and UPR in regulation of skeletal muscle mass in naïve conditions and during cancer cachexia. Our results demonstrate that multiple markers of ER stress are highly activated in skeletal muscle of Lewis lung carcinoma (LLC) and Apc(Min/+) mouse models of cancer cachexia. Treatment of mice with 4-phenylbutyrate (4-PBA), a chemical chaperon and a potent inhibitor of ER stress, significantly reduced skeletal muscle strength and mass in both control and LLC-bearing mice. Blocking the UPR also increased the proportion of fast-type fibers in soleus muscle of both control and LLC-bearing mice. Inhibition of UPR reduced the activity of Akt/mTOR pathway and increased the expression of the components of the ubiquitin-proteasome system and autophagy in LLC-bearing mice. Moreover, we found that the inhibition of UPR causes severe atrophy in cultured myotubes. Our study provides initial evidence that ER stress and UPR pathways are essential for maintaining skeletal muscle mass and strength and for protection against cancer cachexia.-Bohnert, K. R., Gallot, Y. S., Sato, S., Xiong, G., Hindi, S. M., Kumar, A. Inhibition of ER stress and unfolding protein response pathways causes skeletal muscle wasting during cancer cachexia.

A DUF-246 family glycosyltransferase-like gene affects male fertility and the biosynthesis of pectic arabinogalactans.

BACKGROUND: Pectins are a group of structurally complex plant cell wall polysaccharides whose biosynthesis and function remain poorly understood. The pectic polysaccharide rhamnogalacturonan-I (RG-I) has two types of arabinogalactan side chains, type-I and type-II arabinogalactans. To date few enzymes involved in the biosynthesis of pectin have been described. Here we report the identification of a highly conserved putative glycosyltransferase encoding gene, Pectic ArabinoGalactan synthesis-Related (PAGR), affecting the biosynthesis of RG-I arabinogalactans and critical for pollen tube growth. RESULTS: T-DNA insertions in PAGR were identified in Arabidopsis thaliana and were found to segregate at a 1:1 ratio of heterozygotes to wild type. We were unable to isolate homozygous pagr mutants as pagr mutant alleles were not transmitted via pollen. In vitro pollen germination assays revealed reduced rates of pollen tube formation in pollen from pagr heterozygotes. To characterize a loss-of-function phenotype for PAGR, the Nicotiana benthamiana orthologs, NbPAGR-A and B, were transiently silenced using Virus Induced Gene Silencing. NbPAGR-silenced plants exhibited reduced internode and petiole expansion. Cell wall materials from NbPAGR-silenced plants had reduced galactose content compared to the control. Immunological and linkage analyses support that RG-I has reduced type-I arabinogalactan content and reduced branching of the RG-I backbone in NbPAGR-silenced plants. Arabidopsis lines overexpressing PAGR exhibit pleiotropic developmental phenotypes and the loss of apical dominance as well as an increase in RG-I type-II arabinogalactan content. CONCLUSIONS: Together, results support a function for PAGR in the biosynthesis of RG-I arabinogalactans and illustrate the essential roles of these polysaccharides in vegetative and reproductive plant growth.

Comparative transcriptomics reveals patterns of selection in domesticated and wild tomato.

Although applied over extremely short timescales, artificial selection has dramatically altered the form, physiology, and life history of cultivated plants. We have used RNAseq to define both gene sequence and expression divergence between cultivated tomato and five related wild species. Based on sequence differences, we detect footprints of positive selection in over 50 genes. We also document thousands of shifts in gene-expression level, many of which resulted from changes in selection pressure. These rapidly evolving genes are commonly associated with environmental response and stress tolerance. The importance of environmental inputs during evolution of gene expression is further highlighted by large-scale alteration of the light response coexpression network between wild and cultivated accessions. Human manipulation of the genome has heavily impacted the tomato transcriptome through directed admixture and by indirectly favoring nonsynonymous over synonymous substitutions. Taken together, our results shed light on the pervasive effects artificial and natural selection have had on the transcriptomes of tomato and its wild relatives.

A ß-glucuronosyltransferase from Arabidopsis thaliana involved in biosynthesis of type II arabinogalactan has a role in cell elongation during seedling growth.

We have characterized a ß-glucuronosyltransferase (AtGlcAT14A) from Arabidopsis thaliana that is involved in the biosynthesis of type II arabinogalactan (AG). This enzyme belongs to the Carbohydrate Active Enzyme database glycosyltransferase family 14 (GT14). The protein was localized to the Golgi apparatus when transiently expressed in Nicotiana benthamiana. The soluble catalytic domain expressed in Pichia pastoris transferred glucuronic acid (GlcA) to ß-1,6-galactooligosaccharides with degrees of polymerization (DP) ranging from 3-11, and to ß-1,3-galactooligosaccharides of DP5 and 7, indicating that the enzyme is a glucuronosyltransferase that modifies both the ß-1,6- and ß-1,3-galactan present in type II AG. Two allelic T-DNA insertion mutant lines showed 20-35% enhanced cell elongation during seedling growth compared to wild-type. Analyses of AG isolated from the mutants revealed a reduction of GlcA substitution on Gal-ß-1,6-Gal and ß-1,3-Gal, indicating an in vivo role of AtGlcAT14A in synthesis of those structures in type II AG. Moreover, a relative increase in the levels of 3-, 6- and 3,6-linked galactose (Gal) and reduced levels of 3-, 2- and 2,5-linked arabinose (Ara) were seen, suggesting that the mutation in AtGlcAT14A results in a relative increase of the longer and branched ß-1,3- and ß-1,6-galactans. This increase of galactosylation in the mutants is most likely caused by increased availability of the O6 position of Gal, which is a shared acceptor site for AtGlcAT14A and galactosyltransferases in synthesis of type II AG, and thus addition of GlcA may terminate Gal chain extension. We discuss a role for the glucuronosyltransferase in the biosynthesis of type II AG, with a biological role during seedling growth.

O-acetylation of Arabidopsis hemicellulose xyloglucan requires AXY4 or AXY4L, proteins with a TBL and DUF231 domain.

In an Arabidopsis thaliana forward genetic screen aimed at identifying mutants with altered structures of their hemicellulose xyloglucan (axy mutants) using oligosaccharide mass profiling, two nonallelic mutants (axy4-1 and axy4-2) that have a 20 to 35% reduction in xyloglucan O-acetylation were identified. Mapping of the mutation in axy4-1 identified AXY4, a type II transmembrane protein with a Trichome Birefringence-Like domain and a domain of unknown function (DUF231). Loss of AXY4 transcript results in a complete lack of O-acetyl substituents on xyloglucan in several tissues, except seeds. Seed xyloglucan is instead O-acetylated by the paralog AXY4like, as demonstrated by the analysis of the corresponding T-DNA insertional lines. Wall fractionation analysis of axy4 knockout mutants indicated that only a fraction containing xyloglucan is non-O-acetylated. Hence, AXY4/AXY4L is required for the O-acetylation of xyloglucan, and we propose that these proteins represent xyloglucan-specific O-acetyltransferases, although their donor and acceptor substrates have yet to be identified. An Arabidopsis ecotype, Ty-0, has reduced xyloglucan O-acetylation due to mutations in AXY4, demonstrating that O-acetylation of xyloglucan does not impact the plant's fitness in its natural environment. The relationship of AXY4 with another previously identified group of Arabidopsis proteins involved in general wall O-acetylation, reduced wall acetylation, is discussed.

Golgi nucleotide sugar transporter modulates cell wall biosynthesis and plant growth in rice.

Golgi-localized nucleotide sugar transporters (NSTs) are considered essential for the biosynthesis of wall polysaccharides and glycoproteins based on their characteristic transport of a large number of nucleotide sugars to the Golgi lumen. The lack of NST mutants in plants has prevented evaluation of this hypothesis in plants. A previously undescribed Golgi NST mutant, brittle culm14 (bc14), displays reduced mechanical strength caused by decreased cellulose content and altered wall structure, and exhibits abnormalities in plant development. Map-based cloning revealed that all of the observed mutant phenotypes result from a missense mutation in a putative NST gene, Oryza sativa Nucleotide Sugar Transporter1 (OsNST1). OsNST1 was identified as a Golgi-localized transporter by analysis of a fluorescence-tagged OsNST1 expressed in rice protoplast cells and demonstration of UDP-glucose transport activity via uptake assays in yeast. Compositional sugar analyses in total and fractionated wall residues of wild-type and bc14 culms showed a deficiency in the synthesis of glucoconjugated polysaccharides in bc14, indicating that OsNST1 supplies the glucosyl substrate for the formation of matrix polysaccharides, and thereby modulates cellulose biosynthesis. OsNST1 is ubiquitously expressed, with high expression in mechanical tissues. The inferior mechanical strength and abnormal development of bc14 plants suggest that OsNST1 has pleiotropic effects on cell wall biosynthesis and plant growth. Identification of OsNST1 has improved our understanding of how cell wall polysaccharide synthesis is regulated by Golgi NSTs in plants.

Hemicellulose biosynthesis.

One major component of plant cell walls is a diverse group of polysaccharides, the hemicelluloses. Hemicelluloses constitute roughly one-third of the wall biomass and encompass the heteromannans, xyloglucan, heteroxylans, and mixed-linkage glucan. The fine structure of these polysaccharides, particularly their substitution, varies depending on the plant species and tissue type. The hemicelluloses are used in numerous industrial applications such as food additives as well as in medicinal applications. Their abundance in lignocellulosic feedstocks should not be overlooked, if the utilization of this renewable resource for fuels and other commodity chemicals becomes a reality. Fortunately, our understanding of the biosynthesis of the various hemicelluloses in the plant has increased enormously in recent years mainly through genetic approaches. Taking advantage of this knowledge has led to plant mutants with altered hemicellulosic structures demonstrating the importance of the hemicelluloses in plant growth and development. However, while we are on a solid trajectory in identifying all necessary genes/proteins involved in hemicellulose biosynthesis, future research is required to combine these single components and assemble them to gain a holistic mechanistic understanding of the biosynthesis of this important class of plant cell wall polysaccharides.

The rice dynamin-related protein DRP2B mediates membrane trafficking, and thereby plays a critical role in secondary cell wall cellulose biosynthesis.

Membrane trafficking between the plasma membrane (PM) and intracellular compartments is an important process that regulates the deposition and metabolism of cell wall polysaccharides. Dynamin-related proteins (DRPs), which function in membrane tubulation and vesiculation are closely associated with cell wall biogenesis. However, the molecular mechanisms by which DRPs participate in cell wall formation are poorly understood. Here, we report the functional characterization of Brittle Culm3 (BC3), a gene encoding OsDRP2B. Consistent with the expression of BC3 in mechanical tissues, the bc3 mutation reduces mechanical strength, which results from decreased cellulose content and altered secondary wall structure. OsDRP2B, one of three members of the DRP2 subfamily in rice (Oryza sativa L.), was identified as an authentic membrane-associated dynamin via in vitro biochemical analyses. Subcellular localization of fluorescence-tagged OsDRP2B and several compartment markers in protoplast cells showed that this protein not only lies at the PM and the clathrin-mediated vesicles, but also is targeted to the trans-Golgi network (TGN). An FM4-64 uptake assay in transgenic plants that express green fluorescent protein-tagged OsDRP2B verified its involvement in an endocytic pathway. BC3 mutation and overexpression altered the abundance of cellulose synthase catalytic subunit 4 (OsCESA4) in the PM and in the endomembrane systems. All of these findings lead us to conclude that OsDRP2B participates in the endocytic pathway, probably as well as in post-Golgi membrane trafficking. Mutation of OsDRP2B disturbs the membrane trafficking that is essential for normal cellulose biosynthesis of the secondary cell wall, thereby leading to inferior mechanical properties in rice plants.

Arabidopsis CSLD1 and CSLD4 are required for cellulose deposition and normal growth of pollen tubes.

The cell wall is important for pollen tube growth, but little is known about the molecular mechanism that controls cell wall deposition in pollen tubes. Here, the functional characterization of the pollen-expressed Arabidopsis cellulose synthase-like D genes CSLD1 and CSLD4 that are required for pollen tube growth is reported. Both CSLD1 and CSLD4 are highly expressed in mature pollen grains and pollen tubes. The CSLD1 and CSLD4 proteins are located in the Golgi apparatus and transported to the plasma membrane of the tip region of growing pollen tubes, where cellulose is actively synthesized. Mutations in CSLD1 and CSLD4 caused a significant reduction in cellulose deposition in the pollen tube wall and a remarkable disorganization of the pollen tube wall layers, which disrupted the genetic transmission of the male gametophyte. In csld1 and csld4 single mutants and in the csld1 csld4 double mutant, all the mutant pollen tubes exhibited similar phenotypes: the pollen tubes grew extremely abnormally both in vitro and in vivo, which indicates that CSLD1 and CSLD4 are not functionally redundant. Taken together, these results suggest that CSLD1 and CSLD4 play important roles in pollen tube growth, probably through participation in cellulose synthesis of the pollen tube wall.

Rice cellulose synthase-like D4 is essential for normal cell-wall biosynthesis and plant growth.

Cellulose synthase-like (CSL) proteins of glycosyltransferase family 2 (GT2) are believed to be involved in the biosynthesis of cell-wall polymers. The CSL D sub-family (CSLD) is common to all plants, but the functions of CSLDs remain to be elucidated. We report here an in-depth characterization of a narrow leaf and dwarf1 (nd1) rice mutant that shows significant reduction in plant growth due to retarded cell division. Map-based cloning revealed that ND1 encodes OsCSLD4, one of five members of the CSLD sub-family in rice. OsCSLD4 is mainly expressed in tissues undergoing rapid growth. Expression of OsCSLD4 fluorescently tagged at the C- or N-terminus in rice protoplast cells or Nicotiana benthamiana leaves showed that the protein is located in the endoplasmic reticulum or Golgi vesicles. Golgi localization was verified using phenotype-rescued transgenic plants expressing OsCSLD4-GUS under the control of its own promoter. Two phenotype-altered tissues, culms and root tips, were used to investigate the specific wall defects. Immunological studies and monosaccharide compositional and glycosyl linkage analyses explored several wall compositional effects caused by disruption of OsCSLD4, including alterations in the structure of arabinoxylan and the content of cellulose and homogalacturonan, which are distinct in the monocot grass species Oryza sativa (rice). The inconsistent alterations in the two tissues and the observable structural defects in primary walls indicate that OsCSLD4 plays important roles in cell-wall formation and plant growth.

A missense mutation in the transmembrane domain of CESA4 affects protein abundance in the plasma membrane and results in abnormal cell wall biosynthesis in rice.

Cellulose synthase (CESA) is a critical catalytic subunit of the cellulose synthase complex responsible for glucan chain elongation. Our knowledge about how CESA functions is still very limited. Here, we report the functional characterization of a rice mutant, brittle culm11, that shows growth retardation and dramatically reduced plant strength. Map-based cloning revealed that all the mutant phenotypes result from a missense mutation in OsCESA4 (G858R), a highly conserved residue at the end of the fifth transmembrane domain. The aberrant secondary cell wall of the mutant plants is attributed to significantly reduced cellulose content, abnormal secondary wall structure of sclerenchyma cells, and overall altered wall composition, as detected by chemical analyses and immunochemical staining. Importantly, we have found that this point mutation decreases the abundance of OsCESA4 in the plasma membrane, probably due to a defect in the process of CESA complex secretion. The data from our biochemical, genetic, and pharmacological analyses indicate that this residue is critical for maintaining the normal level of CESA proteins in the plasma membrane.

Canonical NF-κB signaling regulates satellite stem cell homeostasis and function during regenerative myogenesis.

Skeletal muscle regeneration in adults is attributed to the presence of satellite stem cells that proliferate, differentiate, and eventually fuse with injured myofibers. However, the signaling mechanisms that regulate satellite cell homeostasis and function remain less understood. While IKKß-mediated canonical NF-κB signaling has been implicated in the regulation of myogenesis and skeletal muscle mass, its role in the regulation of satellite cell function during muscle regeneration has not been fully elucidated. Here, we report that canonical NF-κB signaling is induced in skeletal muscle upon injury. Satellite cell-specific inducible ablation of IKKß attenuates skeletal muscle regeneration in adult mice. Targeted ablation of IKKß also reduces the number of satellite cells in injured skeletal muscle of adult mice, potentially through inhibiting their proliferation and survival. We also demonstrate that the inhibition of specific components of the canonical NF-κB pathway causes precocious differentiation of cultured satellite cells both ex vivo and in vitro. Finally, our results highlight that the constitutive activation of canonical NF-κB signaling in satellite cells also attenuates skeletal muscle regeneration following injury in adult mice. Collectively, our study demonstrates that the proper regulation of canonical NF-κB signaling is important for the regeneration of adult skeletal muscle.

The Toll-Like Receptor/MyD88/XBP1 Signaling Axis Mediates Skeletal Muscle Wasting during Cancer Cachexia.

Skeletal muscle wasting causes both morbidity and mortality of cancer patients. Accumulating evidence suggests that the markers of endoplasmic reticulum (ER) stress and unfolded protein response (UPR) pathways are increased in skeletal muscle under multiple catabolic conditions, including cancer. However, the signaling mechanisms and the role of individual arms of the UPR in the regulation of skeletal muscle mass remain largely unknown. In the present study, we demonstrated that gene expression of Toll-like receptors (TLRs) and myeloid differentiation primary response gene 88 (MyD88) was increased in skeletal muscle in a Lewis lung carcinoma (LLC) model of cancer cachexia. Targeted ablation of MyD88 inhibits the loss of skeletal muscle mass and strength in LLC tumor-bearing mice. Inhibition of MyD88 attenuates the LLC-induced activation of the UPR in skeletal muscle of mice. Moreover, muscle-specific deletion of X-box binding protein 1 (XBP1), a major downstream target of IRE1α arm of the UPR, ameliorates muscle wasting in LLC tumor-bearing mice. Our results also demonstrate that overexpression of an active form of XBP1 caused atrophy in cultured myotubes. In contrast, knockdown of XBP1 inhibits myotube atrophy in response to LLC or C26 adenocarcinoma cell conditioned medium. Collectively, our results demonstrate that TLR/MyD88-mediated activation of XBP1 causes skeletal muscle wasting in LLC tumor-bearing mice.

Growth- and stress-related defects associated with wall hypoacetylation are strigolactone-dependent.

Mutants affected in the Arabidopsis TBL29/ESK1 xylan O-acetyltransferase display a strong reduction in total wall O-acetylation accompanied by a dwarfed plant stature, collapsed xylem morphology, and enhanced freezing tolerance. A newly identified tbl29/esk1 suppressor mutation reduces the expression of the MAX4 gene, affecting the biosynthesis of methyl carlactonoate (MeCLA), an active strigolactone (SL). Genetic and biochemical evidence suggests that blocking the biosynthesis of this SL is sufficient to recover all developmental and stress-related defects associated with the TBL29/ESK1 loss of function without affecting its direct effect-reduced wall O-acetylation. Altered levels of the MAX4 SL biosynthetic gene, reduced branch number, and higher levels of MeCLA, were also found in tbl29/esk1 plants consistent with a constitutive activation of the SL pathway. These results suggest that the reduction in O-acetyl substituents in xylan is not directly responsible for the observed tbl29/esk1 phenotypes. Alternatively, plants may perceive defects in the structure of wall polymers and/or wall architecture activating the SL hormonal pathway as a compensatory mechanism.

TAK1 regulates skeletal muscle mass and mitochondrial function.

Skeletal muscle mass is regulated by a complex array of signaling pathways. TGF-ß-activated kinase 1 (TAK1) is an important signaling protein, which regulates context-dependent activation of multiple intracellular pathways. However, the role of TAK1 in the regulation of skeletal muscle mass remains unknown. Here, we report that inducible inactivation of TAK1 causes severe muscle wasting, leading to kyphosis, in both young and adult mice.. Inactivation of TAK1 inhibits protein synthesis and induces proteolysis, potentially through upregulating the activity of the ubiquitin-proteasome system and autophagy. Phosphorylation and enzymatic activity of AMPK are increased, whereas levels of phosphorylated mTOR and p38 MAPK are diminished upon inducible inactivation of TAK1 in skeletal muscle. In addition, targeted inactivation of TAK1 leads to the accumulation of dysfunctional mitochondria and oxidative stress in skeletal muscle of adult mice. Inhibition of TAK1 does not attenuate denervation-induced muscle wasting in adult mice. Finally, TAK1 activity is highly upregulated during overload-induced skeletal muscle growth, and inactivation of TAK1 prevents myofiber hypertrophy in response to functional overload. Overall, our study demonstrates that TAK1 is a key regulator of skeletal muscle mass and oxidative metabolism.