23 research outputs found
Expression and regulation of type 2A protein phosphatases and alpha4 signalling in cardiac health and hypertrophy
Abstract Cardiac physiology and hypertrophy are regulated
by the phosphorylation status of many proteins, which
is partly controlled by a poorly defined type 2A protein
phosphatase-alpha4 intracellular signalling axis. Quantitative
PCR analysis revealed that mRNA levels of the type
2A catalytic subunits were differentially expressed in H9c2
cardiomyocytes (PP2ACb[PP2ACa[PP4C[PP6C),
NRVM (PP2ACb[PP2ACa = PP4C = PP6C), and
adult rat ventricular myocytes (PP2ACa[
PP2ACb[PP6C[PP4C). Western analysis confirmed
that all type 2A catalytic subunits were expressed in H9c2
cardiomyocytes; however, PP4C protein was absent in
adult myocytes and only detectable following 26S proteasome
inhibition. Short-term knockdown of alpha4 protein
expression attenuated expression of all type 2A catalytic
subunits. Pressure overload-induced left ventricular (LV)
hypertrophy was associated with an increase in both
PP2AC and alpha4 protein expression. Although PP6C
expression was unchanged, expression of PP6C regulatory
subunits (1) Sit4-associated protein 1 (SAP1) and (2)
ankyrin repeat domain (ANKRD) 28 and 44 proteins was
elevated, whereas SAP2 expression was reduced in
hypertrophied LV tissue. Co-immunoprecipitation studies
demonstrated that the interaction between alpha4 and
PP2AC or PP6C subunits was either unchanged or reduced
in hypertrophied LV tissue, respectively. Phosphorylation
status of phospholemman (Ser63 and Ser68) was significantly
increased by knockdown of PP2ACa, PP2ACb, or
PP4C protein expression. DNA damage assessed by histone
H2A.X phosphorylation (cH2A.X) in hypertrophied tissue
remained unchanged. However, exposure of cardiomyocytes
to H2O2 increased levels of cH2A.X which was
unaffected by knockdown of PP6C expression, but was
abolished by the short-term knockdown of alpha4 expression.
This study illustrates the significance and altered
activity of the type 2A protein phosphatase-alpha4 complex
in healthy and hypertrophied myocardium
Essential Roles of the Tap42-Regulated Protein Phosphatase 2A (PP2A) Family in Wing Imaginal Disc Development of Drosophila melanogaster
Protein ser/thr phosphatase 2A family members (PP2A, PP4, and PP6) are implicated in the control of numerous biological processes, but our understanding of the in vivo function and regulation of these enzymes is limited. In this study, we investigated the role of Tap42, a common regulatory subunit for all three PP2A family members, in the development of Drosophila melanogaster wing imaginal discs. RNAi-mediated silencing of Tap42 using the binary Gal4/UAS system and two disc drivers, pnr- and ap-Gal4, not only decreased survival rates but also hampered the development of wing discs, resulting in a remarkable thorax cleft and defective wings in adults. Silencing of Tap42 also altered multiple signaling pathways (HH, JNK and DPP) and triggered apoptosis in wing imaginal discs. The Tap42RNAi-induced defects were the direct result of loss of regulation of Drosophila PP2A family members (MTS, PP4, and PPV), as enforced expression of wild type Tap42, but not a phosphatase binding defective Tap42 mutant, rescued fly survivorship and defects. The experimental platform described herein identifies crucial roles for Tap42β’phosphatase complexes in governing imaginal disc and fly development
NMR Studies of the C-Terminus of alpha4 Reveal Possible Mechanism of Its Interaction with MID1 and Protein Phosphatase 2A
Alpha4 is a regulatory subunit of the protein phosphatase family of enzymes and plays an essential role in regulating the catalytic subunit of PP2A (PP2Ac) within the rapamycin-sensitive signaling pathway. Alpha4 also interacts with MID1, a microtubule-associated ubiquitin E3 ligase that appears to regulate the function of PP2A. The C-terminal region of alpha4 plays a key role in the binding interaction of PP2Ac and MID1. Here we report on the solution structure of a 45-amino acid region derived from the C-terminus of alpha4 (alpha45) that binds tightly to MID1. In aqueous solution, alpha45 has properties of an intrinsically unstructured peptide although chemical shift index and dihedral angle estimation based on chemical shifts of backbone atoms indicate the presence of a transient Ξ±-helix. Alpha45 adopts a helix-turn-helix HEAT-like structure in 1% SDS micelles, which may mimic a negatively charged surface for which alpha45 could bind. Alpha45 binds tightly to the Bbox1 domain of MID1 in aqueous solution and adopts a structure consistent with the helix-turn-helix structure observed in 1% SDS. The structure of alpha45 reveals two distinct surfaces, one that can interact with a negatively charged surface, which is present on PP2A, and one that interacts with the Bbox1 domain of MID1
AACR Project GENIE: 100,000 cases and beyond.
The American Association for Cancer Research (AACR) Project GENIE is an international pan-cancer registry with the goal to inform cancer research and clinical care worldwide. Founded in late 2015, the milestone GENIE 9.1-public release contains data from \u3e110,000 tumors from \u3e100,000 people treated at 19 cancer centers from USA, Canada, the United Kingdom, France, Netherlands, and Spain. Here, we demonstrate use of these real-world data, harmonized through a centralized data resource to accurately predict enrollment on genome-guided trials, discover driver alterations in rare tumors, and identify cancer types without actionable mutations that could benefit from comprehensive genomic analysis. The extensible data infrastructure and governance framework support additional deep patient phenotyping through biopharma collaborations, and expansion to include new data types such as cell-free DNA sequencing. AACR Project GENIE continues to serve a global precision medicine knowledgebase of increasing impact to inform clinical decision making and bring together cancer researchers internationally
Discovery of new substrates of the elongation factor-2 kinase suggests a broader role in the cellular nutrient response
Elongation Factor-2 Kinase (eEF2K) in an unusual mammalian enzyme that has one known substrate, elongation factor-2. It belongs to a class of kinases, called alpha kinases, that has little sequence identity to the > 500 conventional protein kinases, but performs the same reaction and has similar catalytic residues. The phosphorylation of eEF2 blocks translation elongation, which is thought to be critical to regulating cellular energy usage. Here we report a system for discovering new substrates of alpha kinases and identify the first new substrates of eEF2K including AMPK and alpha4, and determine a sequence motif for the kinase that shows a requirement for threonine residues as the target of phosphorylation. These new substrates suggest that eEF2K has a more diverse role in regulating cellular energy usage that involves multiple pathways and regulatory feedback
Supplementary Tables from Genomic Characterization and Clinical Outcomes of Patients with Peritoneal Metastases from the AACR GENIE Biopharma Collaborative Colorectal Cancer Registry
Supplementary Tables 1-9</p
FIGURE 2 from Genomic Characterization and Clinical Outcomes of Patients with Peritoneal Metastases from the AACR GENIE Biopharma Collaborative Colorectal Cancer Registry
OS from diagnosis of advanced disease (A, PM vs. non-PM; B, PM only vs. PM and other sites vs. non-PM; C, stage IV: PM vs. non-PM; D, stage IβIII with later development of distant metastasis: PM vs. non-PM).</p
TABLE 3 from Genomic Characterization and Clinical Outcomes of Patients with Peritoneal Metastases from the AACR GENIE Biopharma Collaborative Colorectal Cancer Registry
Frequencies of genomic alterations in tumor samplesβResults are displayed for genomic alterations present in at least 7% of tumor specimens and tested in at least 20% of the tumor specimens, as well as NRAS and MED12. Frequencies do not reflect OncoKB annotation. Details of specific NGS panels performed can be found in the GENIE data guide (www.aacr.org/wp-content/uploads/2022/02/GENIE_data_guide_11.0-public-1.pdf)</p
Supplementary Figure 1 from Genomic Characterization and Clinical Outcomes of Patients with Peritoneal Metastases from the AACR GENIE Biopharma Collaborative Colorectal Cancer Registry
Definitions of patient-level MSI and MMR statuses</p