Validation of GCN5L1/BLOC1S1/BLOS1 antibodies using knockout cells and tissue.

GCN5L1, also known as BLOC1S1 and BLOS1, is a small intracellular protein involved in many key biological processes. Over the last decade, GCN5L1 has been implicated in the regulation of protein lysine acetylation, energy metabolism, endo-lysosomal function, and cellular immune pathways. An increasing number of published papers have used commercially-available reagents to interrogate GCN5L1 function. However, in many cases these reagents have not been rigorously validated, leading to potentially misleading results. In this report we tested several commercially-available antibodies for GCN5L1, and found that two-thirds of those available did not unambiguously detect the protein by western blot in cultured mouse cells or ex vivo liver tissue. These data suggest that previously published studies which used these unverified antibodies to measure GCN5L1 protein abundance, in the absence of other independent methods of corroboration, should be interpreted with appropriate caution.

Renal surgery in patients with a duplicated inferior vena cava: a case series and review of the literature.

Abnormal inferior vena cava (IVC) anatomy may present unique challenges for urologists when performing retroperitoneal surgery. Duplication of the IVC is one such anomalous variation and can be found in up to 3% of the population. Misunderstanding of the implications of this aberrant anatomy may lead to intraoperative or postoperative complications. Here, we present two cases of patients undergoing renal surgeries with duplicate IVC. We then review the embryologic origin and anatomic findings in those with abnormal IVC anatomy as well as discuss the surgical implications and considerations for urologists.

The Risk of Prostate Cancer Progression in Active Surveillance Patients with Bilateral Disease Detected by Combined Magnetic Resonance Imaging-Fusion and Systematic Biopsy.

PURPOSE: We sought to evaluate whether bilateral prostate cancer detected at active surveillance (AS) enrollment is associated with progression to Grade Group (GG) ≥2 and to compare the efficacy of combined targeted biopsy plus systematic biopsy (Cbx) vs systematic biopsy (Sbx) or targeted biopsy alone to detect bilateral disease. MATERIALS AND METHODS: A prospectively maintained database of patients referred to our institution from 2007-2020 was queried. The study cohort included all AS patients with GG1 on confirmatory Cbx and followup of at least 1 year. Cox proportional hazard analysis identified baseline characteristics associated with progression to ≥GG2 at any point throughout followup. RESULTS: Of 579 patients referred, 103 patients had GG1 on Cbx and were included in the study; 49/103 (47.6%) patients progressed to ≥GG2, with 30/72 (41.7%) patients with unilateral disease progressing and 19/31 (61.3%) patients with bilateral disease progressing. Median time to progression was 68 months vs 52 months for unilateral and bilateral disease, respectively (p=0.006). Both prostate specific antigen density (HR 1.72, p=0.005) and presence of bilateral disease (HR 2.21, p=0.012) on confirmatory biopsy were associated with AS progression. At time of progression, GG and risk group were significantly higher in patients with bilateral versus unilateral disease. Cbx detected 16% more patients with bilateral disease than Sbx alone. CONCLUSIONS: Bilateral disease and prostate specific antigen density at confirmatory Cbx conferred greater risk of earlier AS progression. Cbx was superior to Sbx for identifying bilateral disease. AS risk-stratification protocols may benefit from including presence of bilateral disease and should use Cbx to detect bilateral disease.

Expansion of the Family of Molecular Nanoparticles of Cerium Dioxide and Their Catalytic Scavenging of Hydroxyl Radicals.

The syntheses, crystal structures, and catalytic radical scavenging activity are reported for four new molecular clusters that have resulted from a bottom-up molecular approach to nanoscale CeO2. They are [Ce6O4(OH)4(dmb)12(H2O)4] (dmb- = 2,6-dimethoxybenzoate), [Ce16O17(OH)6(O2CPh)24(HO2CPh)4], [Ce19O18(OH)9(O2CPh)27(H2O)(py)3], and [Ce24O27(OH)9(O2CPh)30(py)4]. They represent a major expansion of our family of so-called "molecular nanoparticles" of this metal oxide to seven members, and their crystal structures confirm that their cores all possess the fluorite structure of bulk CeO2. In addition, they have allowed the identification of surface features such as the close location of multiple Ce3+ ions and organic ligand binding modes not seen previously. The ability of all seven members to catalytically scavenge reactive oxygen species has been investigated using HO• radicals, an important test reaction in the ceria nanoparticle biomedical literature, and most have been found to exhibit excellent antioxidant activities compared to traditional ceria nanoparticles, with their activity correlating inversely with their surface Ce3+ content.

Truly Monodisperse Molecular Nanoparticles of Cerium Dioxide of 2.4†nm dimensions: A {Ce100 O167 } Cluster.

Ultra-small nanoparticles of CeO2 obtained in molecular form, so-called molecular nanoparticles, have been limited to date to a family whose largest member is of nuclearity Ce40 with a {Ce40 O58 } core atom count. Herein we report that a synthetic procedure has been developed to the cation [Ce100 O149 (OH)18 (O2 CPh)60 (PhCO2 H)12 (H2 O)20 ]16+ , a member with a much higher Ce100 nuclearity and a {Ce100 O167 } core that is more akin to the smallest ceria nanoparticles. Its crystal structure reveals it to possess a 2.4 nm size and high D2d symmetry, and it has also allowed identification of core surface features including facet composition, the presence and location of Ce3+ and H+ (i.e. HO- ) ions, and the binding modes of the ligand monolayer of benzoate, benzoic acid, and water ligands.

GCN5-like protein 1 (GCN5L1) controls mitochondrial content through coordinated regulation of mitochondrial biogenesis and mitophagy.

Cellular mitochondrial content is governed by the competing processes of organelle biogenesis and degradation. It is proposed that these programs are tightly regulated to ensure that the cell maintains sufficient organelles to meet its biosynthetic, energetic, and other homeostatic requirements. We recently reported that GCN5L1, a putative nutrient-sensing regulator, controls mitochondrial removal by autophagy. Here we show that genetic deletion of GCN5L1 has a direct positive effect on the expression and activity of Transcriptional Factor EB (TFEB), which acts as a master regulator of autophagy. Surprisingly, the induction of TFEB-mediated autophagy pathways does not diminish cellular mitochondrial content, as its activity is countered by induction of the mitochondrial biogenesis transcriptional co-activator PPARγ coactivator 1α (PGC-1α). Concurrent induction of the TFEB and PGC-1α pathways results in an increased mitochondrial turnover rate in GCN5L1(-/-) cells. Finally, we show that genetic knockdown of either TFEB or PGC-1α leads to a corresponding decrease in the expression of the other gene, indicating that these proteins act coordinately, and in opposition, to maintain cellular mitochondrial content in response to the modulation of nutrient-sensing signatures.

Regulation of autophagy and mitophagy by nutrient availability and acetylation.

Normal cellular function is dependent on a number of highly regulated homeostatic mechanisms, which act in concert to maintain conditions suitable for life. During periods of nutritional deficit, cells initiate a number of recycling programs which break down complex intracellular structures, thus allowing them to utilize the energy stored within. These recycling systems, broadly named "autophagy", enable the cell to maintain the flow of nutritional substrates until they can be replenished from external sources. Recent research has shown that a number of regulatory components of the autophagy program are controlled by lysine acetylation. Lysine acetylation is a reversible post-translational modification that can alter the activity of enzymes in a number of cellular compartments. Strikingly, the main substrate for this modification is a product of cellular energy metabolism: acetyl-CoA. This suggests a direct and intricate link between fuel metabolites and the systems which regulate nutritional homeostasis. In this review, we examine how acetylation regulates the systems that control cellular autophagy, and how global protein acetylation status may act as a trigger for recycling of cellular components in a nutrient-dependent fashion. In particular, we focus on how acetylation may control the degradation and turnover of mitochondria, the major source of fuel-derived acetyl-CoA.

Restricted mitochondrial protein acetylation initiates mitochondrial autophagy.

Because nutrient-sensing nuclear and cytosolic acetylation mediates cellular autophagy, we investigated whether mitochondrial acetylation modulates mitochondrial autophagy (mitophagy). Knockdown of GCN5L1, a component of the mitochondrial acetyltransferase machinery, diminished mitochondrial protein acetylation and augmented mitochondrial enrichment of autophagy mediators. This program was disrupted by SIRT3 knockdown. Chronic GCN5L1 depletion increased mitochondrial turnover and reduced mitochondrial protein content and/or mass. In parallel, mitochondria showed blunted respiration and enhanced 'stress-resilience'. Genetic disruption of autophagy mediators Atg5 and p62 (also known as SQSTM1), as well as GCN5L1 reconstitution, abolished deacetylation-induced mitochondrial autophagy. Interestingly, this program is independent of the mitophagy E3-ligase Parkin (also known as PARK2). Taken together, these data suggest that deacetylation of mitochondrial proteins initiates mitochondrial autophagy in a canonical autophagy-mediator-dependent program and shows that modulation of this regulatory program has ameliorative mitochondrial homeostatic effects.

Molecular Characterization of Metastatic Oncocytoma With Exceptional Response to Treatment: A Case Report.

Comprehensive molecular characterization and effective therapy in a rare case of metastatic renal oncocytoma.

Phosphoinositide-dependent kinase-1 and protein kinase Cδ contribute to endothelin-1 constriction and elevated blood pressure in intermittent hypoxia.

Obstructive sleep apnea (OSA) is associated with cardiovascular complications including hypertension. Previous findings from our laboratory indicate that exposure to intermittent hypoxia (IH), to mimic sleep apnea, increases blood pressure in rats. IH also increases endothelin-1 (ET-1) constrictor sensitivity in a protein kinase C (PKC) δ-dependent manner in mesenteric arteries. Because phosphoinositide-dependent kinase-1 (PDK-1) regulates PKCδ activity, we hypothesized that PDK-1 contributes to the augmented ET-1 constrictor sensitivity and elevated blood pressure following IH. Male Sprague-Dawley rats were exposed to either sham or IH (cycles between 21% O(2)/0% CO(2) and 5% O(2)/5% CO(2)) conditions for 7 h/day for 14 or 21 days. The contribution of PKCδ and PDK-1 to ET-1-mediated vasoconstriction was assessed in mesenteric arteries using pharmacological inhibitors. Constrictor sensitivity to ET-1 was enhanced in arteries from IH-exposed rats. Inhibition of PKCδ or PDK-1 blunted ET-1 constriction in arteries from IH but not sham group rats. Western analysis revealed similar levels of total and phosphorylated PDK-1 in arteries from sham and IH group rats but decreased protein-protein interaction between PKCδ and PDK-1 in arteries from IH- compared with sham-exposed rats. Blood pressure was increased in rats exposed to IH, and treatment with the PDK-1 inhibitor OSU-03012 [2-amino-N-{4-[5-(2-phenanthrenyl)-3-(trifluoromethyl)-1H-pyrazol-1-yl]-phenyl}-acetamide] (33 mg/day) lowered blood pressure in IH but not sham group rats. Our results suggest that exposure to IH unmasks a role for PDK-1 in regulating ET-1 constrictor sensitivity and blood pressure that is not present under normal conditions. These novel findings suggest that PDK-1 may be a uniquely effective antihypertensive therapy for OSA patients.

Identification of a molecular component of the mitochondrial acetyltransferase programme: a novel role for GCN5L1.

SIRT3 (sirtuin 3) modulates respiration via the deacetylation of lysine residues in electron transport chain proteins. Whether mitochondrial protein acetylation is controlled by a counter-regulatory program has remained elusive. In the present study we identify an essential component of this previously undefined mitochondrial acetyltransferase system. We show that GCN5L1 [GCN5 (general control of amino acid synthesis 5)-like 1; also known as Bloc1s1] counters the acetylation and respiratory effects of SIRT3. GCN5L1 is mitochondrial-enriched and displays significant homology with a prokaryotic acetyltransferase. Genetic knockdown of GCN5L1 blunts mitochondrial protein acetylation, and its reconstitution in intact mitochondria restores protein acetylation. GCN5L1 interacts with and promotes acetylation of SIRT3 respiratory chain targets and reverses global SIRT3 effects on mitochondrial protein acetylation, respiration and bioenergetics. The results of the present study identify GCN5L1 as a critical prokaryote-derived component of the mitochondrial acetyltransferase programme.

Tumorigenesis Mechanisms Found in Hereditary Renal Cell Carcinoma: A Review.

Renal cell carcinoma is a heterogenous cancer composed of an increasing number of unique subtypes each with their own cellular and tumor behavior. The study of hereditary renal cell carcinoma, which composes just 5% of all types of tumor cases, has allowed for the elucidation of subtype-specific tumorigenesis mechanisms that can also be applied to their sporadic counterparts. This review will focus on the major forms of hereditary renal cell carcinoma and the genetic alterations contributing to their tumorigenesis, including von Hippel Lindau syndrome, Hereditary Papillary Renal Cell Carcinoma, Succinate Dehydrogenase-Deficient Renal Cell Carcinoma, Hereditary Leiomyomatosis and Renal Cell Carcinoma, BRCA Associated Protein 1 Tumor Predisposition Syndrome, Tuberous Sclerosis, Birt-Hogg-Dubé Syndrome and Translocation RCC. The mechanisms for tumorigenesis described in this review are beginning to be exploited via the utilization of novel targets to treat renal cell carcinoma in a subtype-specific fashion.

The emerging characterization of lysine residue deacetylation on the modulation of mitochondrial function and cardiovascular biology.

There is emerging recognition of a novel fuel and redox sensing regulatory program that controls cellular adaptation via nonhistone protein lysine residue acetyl posttranslation modifications. This program functions in tissues with high energy demand and oxidative capacity and is highly enriched in the heart. Deacetylation is regulated by NAD(+)-dependent activation of the sirtuin family of proteins, whereas acetyltransferase modifications are controlled by less clearly delineated acetyltransferases. Subcellular localization specific protein targets of lysine-acetyl modification have been identified in the nucleus, cytoplasm, and mitochondria. Despite distinct subcellular localizations, these modifications appear, in large part, to modify mitochondrial properties including respiration, energy production, apoptosis, and antioxidant defenses. These mitochondrial regulatory programs are important in cardiovascular biology, although how protein acetyl modifications effects cardiovascular pathophysiology has not been extensively explored. This review will introduce the role of nonhistone protein lysine residue acetyl modifications, discuss their regulation and biochemistry and present the direct and indirect data implicating their involvement in the heart and vasculature.

Initial Presentation and Outcomes of Testicular Cancer among Different Racial and Ethnic Groups at a Tertiary Care Facility in New Mexico.

INTRODUCTION: This study explored differences in testicular cancer presentation, treatment, compliance and outcomes among ethnicities in New Mexico. METHODS: A retrospective review of patients with testicular cancer treated between 2002 and 2015 was performed. Data included demographics, stage, delays in care, treatments, insurance status and nonadherence rates. RESULTS: Of 186 patients Hispanics and Native Americans presented at higher stage (p <0.01) and delayed treatment (p=0.02). Retroperitoneal lymph node dissection for stage I disease was 28% while stage II was 30%, compared to 18% and 58% nationally, respectively. Of stage II in Hispanic patients 24.5% received retroperitoneal lymph node dissection compared to 41.3% of Caucasians (p <0.05). Regarding chemotherapy Caucasian patients at stage I were more likely than Hispanics to receive chemotherapy (p <0.05). Hispanics had higher rates of nonadherence (p <0.01). Insurance rates did not differ among groups. However, insurance increased the likelihood for receiving chemotherapy/retroperitoneal lymph node dissection only for Caucasians. Lack of insurance increased active surveillance rates for stage I in Hispanics. The incidence of testicular cancer in Hispanics rose by 58% after 2009 (p <0.05). CONCLUSIONS: Minority groups presented at higher stages and delayed treatment. Retroperitoneal lymph node dissection rates differed nationally compared to this cohort with Hispanic patients at higher stage being less likely to receive retroperitoneal lymph node dissection. Meanwhile, Hispanics with stage I are less likely to obtain chemotherapy. Insurance rates did not differ among ethnicities but having insurance did not increase rates of chemotherapy/retroperitoneal lymph node dissection for Hispanics unlike for Caucasians. Meanwhile, lack of insurance increased stage I rates of active surveillance suggesting cultural/financial factors contribute to treatment decisions. Increased health literacy, outreach and access may aid in alleviating these disparities.

Kidney cancer: from genes to therapy.

Renal cell carcinoma incidence is rising worldwide with increasing subtype stratification by the World Health Organization. Each subtype has unique genetic alterations, cell biology changes and clinical findings. Such genetic alterations offer the potential for individualized therapeutic approaches that are rapidly progressing. This review highlights the most common subtypes of renal cell carcinoma, including both hereditary and sporadic forms, with a focus on genetic changes, clinical findings and ongoing clinical trials.

Phosphorus-based ligand effects on the structure and radical scavenging ability of molecular nanoparticles of CeO2.

Two new CeIV/O2- clusters, (pyH)8[Ce10O4(OH)4(O3PPh)12(NO3)12] (1) and [Ce6O4(OH)4(O2PPh2)4(O2CtBu)8] (2), have been prepared that contain P-based ligands for the first time. They were obtained from the reaction of (NH4)2[Ce(NO3)6], PhPO3H2 or Ph2PO2H, and tBuCO2H in a 2 : 1 : 2 molar ratio in pyridine/MeOH (10 : 1 mL). Both compounds contain a {Ce6O4(OH)4} face-capped octahedral core, with 1 containing an additional four CeIV on the outside to give a supertetrahedral Ce10 topology; the {Ce6O8} unit is the smallest recognizable fragment of the fluorite structure of CeO2. The HO˙ radical scavenging activities of 1 and 2 were measured by UV/vis spectral monitoring of methylene blue oxidation by HO˙ radicals in the presence and absence of the Ce/O clusters, and the results compared with those for larger Ce24 and Ce38 molecular nanoparticles of CeO2 prepared in previous work. 1 and 2 are both very poor HO˙ radical scavengers compared with Ce24 and Ce38, a result that is consistent with reports in the literature that PO43- ions inhibit the radical scavenging ability of traditional CeO2 nanoparticles and putatively assigned to PO43- binding to the surface.

Molecular nanoparticles of cerium dioxide: structure-directing effect of halide ions.

The use of halide ions in the synthesis of Ce/O clusters diverts the reaction to two halide-containing products: Cl- gives a new Ce20 nuclearity with both a high 1 : 1 Ce3+ : Ce4+ ratio and a high percentage of (100) facet coverage, whereas F- gives a known Ce6 nuclearity. Both products include bridging halide ions and are thus the first confirmation of non-oxo (OH-/O2-) anion incorporation onto the Ce/O cluster core.

Epstein Barr virus-positive mucocutaneous ulcer of the colon associated Hodgkin lymphoma in Crohn's disease.

Epstein Barr virus (EBV) positive mucocutaneous ulcers (EBVMCU) form part of a spectrum of EBV-associated lymphoproliferative disease. They have been reported in the setting of immunosenescence and iatrogenic immunosuppression, affecting the oropharyngeal mucosa, skin and gastrointestinal tract (GIT). Case reports and series to date suggest a benign natural history responding to conservative management, particularly in the GIT. We report an unusual case of EBVMCU in the colon, arising in the setting of immunosuppression in the treatment of Crohn's disease, with progression to Hodgkin lymphoma 18 mo after cessation of infliximab. The patient presented with multiple areas of segmental colonic ulceration, histologically showing a polymorphous infiltrate with EBV positive Reed-Sternberg-like cells. A diagnosis of EBVMCU was made. The ulcers failed to regress upon cessation of infliximab and methotrexate for 18 mo. Following commencement of prednisolone for her Crohn's disease, the patient developed widespread Hodgkin lymphoma which ultimately presented as a life-threatening lower GIT bleed requiring emergency colectomy. This is the first report of progression of EBVMCU to Hodgkin lymphoma, in the setting of ongoing iatrogenic immunosuppression and inflammatory bowel disease.

The role of sirtuins in modulating redox stressors.

For much of the time since their discovery, the sirtuin family of deacetylase enzymes has been associated with extension of life span. This longevity-promoting capacity in numerous model systems has enabled the sirtuins to gain "celebrity status" in the field of aging research. However, the mechanisms underpinning these changes remain incompletely defined. A general phenotype long associated with aging is the dysregulation of biological systems, which partly occurs via the accumulation of damage over time. One of the major sources of this damage is oxidative stress, which can harm both biological structures and the mechanisms with which they are repaired. It is now becoming clear that the beneficial life-span effects of sirtuins, along with many of their other functions, are closely linked to their ability to regulate systems that control the redox environment. Here we investigate the links between sirtuins and their oxidative/redox environment and review the control mechanisms that are regulated by the activity of sirtuin deacetylase proteins.

Human cytomegalovirus latent infection and associated viral gene expression.

Human cytomegalovirus (HCMV) is a clinically important and ubiquitous herpesvirus. Following primary productive infection the virus is not completely eliminated from the host, but instead establishes a lifelong latent infection without detectable virus production, from where it can reactivate at a later stage to generate new infectious virus. Reactivated HCMV often results in life-threatening disease in immunocompromised individuals, particularly allogeneic stem cell and solid organ transplant recipients, where it remains one of the most difficult opportunistic pathogens that complicate the care of these patients. The ability of HCMV to establish and reactivate from latency is central to its success as a human pathogen, yet latency remains very poorly understood. This article will cover several aspects of HCMV latency, with a focus on current understanding of viral gene expression and functions during this phase of infection.