Abnormal Protein Formation Research Articles

MP04-17 ALTERATIONS IN RNASET2 FACILITATE A PRO TUMOR MICROENVIRONMENT ASSOCIATED WITH WORSE CLINICAL OUTCOMES IN CLEAR CELL RCC

You have accessJournal of UrologyCME1 Apr 2023MP04-17 ALTERATIONS IN RNASET2 FACILITATE A PRO TUMOR MICROENVIRONMENT ASSOCIATED WITH WORSE CLINICAL OUTCOMES IN CLEAR CELL RCC Taylor Peak, Andrew Chang, Nick Chakiryan, Joseph Johnson, Anders Berglund, Doug Marchion, Asmaa Elkenawi, Noel Clark, Natasha Francis, Paola Ramos Echevarria, Nusheen Immen, Jasreman Dhillon, James Mule, Liang Wang, and Brandon Manley Taylor PeakTaylor Peak More articles by this author , Andrew ChangAndrew Chang More articles by this author , Nick ChakiryanNick Chakiryan More articles by this author , Joseph JohnsonJoseph Johnson More articles by this author , Anders BerglundAnders Berglund More articles by this author , Doug MarchionDoug Marchion More articles by this author , Asmaa ElkenawiAsmaa Elkenawi More articles by this author , Noel ClarkNoel Clark More articles by this author , Natasha FrancisNatasha Francis More articles by this author , Paola Ramos EchevarriaPaola Ramos Echevarria More articles by this author , Nusheen ImmenNusheen Immen More articles by this author , Jasreman DhillonJasreman Dhillon More articles by this author , James MuleJames Mule More articles by this author , Liang WangLiang Wang More articles by this author , and Brandon ManleyBrandon Manley More articles by this author View All Author Informationhttps://doi.org/10.1097/JU.0000000000003215.17AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookLinked InTwitterEmail Abstract INTRODUCTION AND OBJECTIVE: M2 macrophage polarization (CD163) leads to T cell exhaustion and worse clinical outcomes in ccRCC patients. Recently, our group has identified a novel splice variant of RNASET2, a protein normally contributing to anti-tumor activity. This splice variant was associated with worse clinical outcomes in ccRCC patients. We set out to determine the expression patterns for M2 macrophages and RNASET2, and their relationship with the RNASET2 splice variant. We hypothesize that the splice variant leads to abnormal protein formation, allowing for a tumor microenvironment that favors growth. METHODS: We obtained primary tumor samples from 75 patients with ccRCC. Tissue sections from these specimens were obtained from the tumor core, tumor edge, and stroma (Figure 1). IHC was performed, staining for RNASET2, M2 macrophage (CD 163), and markers for T cell exhaustion (TIM3, CD8). In addition, RNA sequencing data was utilized to identify the RNASET2 splice variant expression in all tumors. Expression patterns of these cellular markers were analyzed, and then stratified based on whether the tumor was splice variant positive (SV+) or splice variant negative (SV-) RESULTS: Expression of CD163 was inversely correlated with expression of RNASET2 in the tumor core and tumor edge (Figure 2). Interestingly, SV+ tumors had overall higher RNASET2 expression. However, when relative extracellular to intracellular RNASET2 staining was compared, the SV- tumors possessed relatively higher extracellular expression. Finally, when evaluating expression as a prognostic indicator, we found that high co-expression of CD163 and RNASET2 led to worse overall survival relative to low co-expression. This was seen in the tumor (47 vs 138 mo, p<0.001), tumor edge (80 vs 174 mo, p=0.001) and stroma (15 vs 125 mo, p<0.001). CONCLUSIONS: This data represents the first evidence that a relationship exists between M2 macrophages and RNASET2 in kidney cancer. We have demonstrated through staining of patient tissue samples that high RNASET2 expression is correlated with low M2 polarization of macrophages, and vice versa. Furthermore, based on relative extracellular localization, SV- tumors are more effective at secreting RNASET2 to promote anti-tumor effect. Source of Funding: N/A © 2023 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 209Issue Supplement 4April 2023Page: e40 Advertisement Copyright & Permissions© 2023 by American Urological Association Education and Research, Inc.MetricsAuthor Information Taylor Peak More articles by this author Andrew Chang More articles by this author Nick Chakiryan More articles by this author Joseph Johnson More articles by this author Anders Berglund More articles by this author Doug Marchion More articles by this author Asmaa Elkenawi More articles by this author Noel Clark More articles by this author Natasha Francis More articles by this author Paola Ramos Echevarria More articles by this author Nusheen Immen More articles by this author Jasreman Dhillon More articles by this author James Mule More articles by this author Liang Wang More articles by this author Brandon Manley More articles by this author Expand All Advertisement PDF downloadLoading ...

Iron, Neuroinflammation and Neurodegeneration.

Disturbance of the brain homeostasis, either directly via the formation of abnormal proteins or cerebral hypo-perfusion, or indirectly via peripheral inflammation, will activate microglia to synthesise a variety of pro-inflammatory agents which may lead to inflammation and cell death. The pro-inflammatory cytokines will induce changes in the iron proteins responsible for maintaining iron homeostasis, such that increased amounts of iron will be deposited in cells in the brain. The generation of reactive oxygen and nitrogen species, which is directly involved in the inflammatory process, can significantly affect iron metabolism via their interaction with iron-regulatory proteins (IRPs). This underlies the importance of ensuring that iron is maintained in a form that can be kept under control; hence, the elegant mechanisms which have become increasingly well understood for regulating iron homeostasis. Therapeutic approaches to minimise the toxicity of iron include N-acetyl cysteine, non-steroidal anti-inflammatory compounds and iron chelation.

Processing of Mutant β-Amyloid Precursor Protein and the Clinicopathological Features of Familial Alzheimer's Disease.

Alzheimer’s disease (AD) is a complex, multifactorial disease involving many pathological mechanisms. Nonetheless, single pathogenic mutations in amyloid precursor protein (APP) or presenilin 1 or 2 can cause AD with almost all of the clinical and neuropathological features, and therefore, we believe an important mechanism of pathogenesis in AD could be revealed from examining pathogenic APP missense mutations. A comprehensive review of the literature, including clinical, neuropathological, cellular and animal model data, was conducted through PubMed and the databases of Alzforum mutations, HGMD, UniProt, and AD&FTDMDB. Pearson correlation analysis combining the clinical and neuropathological data and aspects of mutant APP processing in cellular models was performed. We find that an increase in Aβ42 has a significant positive correlation with the appearance of neurofibrillary tangles (NFTs) and tends to cause an earlier age of AD onset, while an increase in Aβ40 significantly increases the age at death. The increase in the α-carboxyl terminal fragment (CTF) has a significantly negative correlation with the age of AD onset, and β-CTF has a similar effect without statistical significance. Animal models show that intracellular Aβ is critical for memory defects. Based on these results and the fact that amyloid plaque burden correlates much less well with cognitive impairment than do NFT counts, we propose a “snowball hypothesis”: the accumulation of intraneuronal NFTs caused by extracellular Aβ42 and the increase in intraneuronal APP proteolytic products (CTFs and Aβs) could cause cellular organelle stress that leads to neurodegeneration in AD, which then resembles the formation of abnormal protein “snowballs” both inside and outside of neurons.

Clinical characteristics of laryngeal versus nonlaryngeal amyloidosis.

Amyloidosis represents a heterogeneous group of disorders marked by abnormal protein formation and deposition. Laryngeal amyloidosis is rare and classically thought to remain isolated with little risk of systemic involvement or associated malignancy. This study sought to further characterize differences in clinical characteristics between patients with laryngeal and nonlaryngeal amyloidosis. Retrospective case-control study. The Stanford Translational Research Integrated Database Environment was searched to identify patients with biopsy-confirmed laryngeal amyloidosis and patients with amyloidosis without laryngeal involvement on endoscopy. Mann-Whitney U and χ2 tests were used for statistical analysis. Of 865 patients treated for amyloidosis between 1996 and 2016, 22 (2.5%) patients with biopsy-proven laryngeal amyloidosis were identified. An additional 22 patients with amyloidosis of a different organ and negative laryngoscopy-and therefore without laryngeal amyloidosis-were identified as controls. Compared to these controls with nonlaryngeal amyloidosis, patients with laryngeal amyloidosis were younger (mean age 52.8 years vs. 68.4 years, P < .0006), and 18% had additional organ involvement. Immunoglobulin light-chain amyloidosis was the most common subtype in both groups of patients. Eighty-six percent of patients with laryngeal amyloidosis required surgical excision, and of these patients, over 30% required multiple excisions. There is a significant rate (18%) of multiorgan involvement in patients with laryngeal amyloidosis, which contradicts conventional concepts that this is an isolated disorder. This finding could have a significant impact on the evaluation and management of patients with laryngeal amyloidosis. 3b. Laryngoscope, 128:670-674, 2018.

Editorial: Genetics and epigenetics of fetal alcohol spectrum disorders.

Editorial: Genetics and epigenetics of fetal alcohol spectrum disorders.

Identification of a novel type of small molecule inhibitor against HIV-1.

Here we report a new chemical inhibitor against HIV-1 with a novel structure and mode of action. The inhibitor, designated as A1836, inhibited HIV-1 replication and virus production with a 50% inhibitory concentration (IC50) of 2.0 μM in an MT-4 cell-based and cytopathic protection antiviral assay, while its 50% cytotoxic concentration (CC50) was much higher than 50 μM. Examination of the effect of A1836 on in vitro HIV-1 reverse transcriptase (RT) and integrase showed that neither were molecular targets of A1836. The characterization and re-infection assay of the HIV-1 virions generated in the presence of A1836 showed that the synthesis of early RT products in the cells infected with the virions was inhibited dose-dependently, due in part to abnormal protein formation within the virions, thus resulting in an impaired infectivity. These results suggest that A1836 might be a novel candidate for the development of a new type of HIV-1 inhibitor. [BMB Reports 2015; 48(2): 121-126]

Expression and activity of l-isoaspartyl methyltransferase decrease in stage progression of human astrocytic tumors

Protein l-isoaspartyl methyltransferase (PIMT) functions as a repair enzyme that acts upon damaged proteins bearing abnormal aspartyl residues. We previously reported that PIMT expression and activity are reduced by half in human epileptic hippocampus. Here we investigated PIMT regulation in astrocytic tumors, which are the most common human brain tumors. PIMT expression and enzyme activity were significantly decreased in all grades of human astrocytic tumors. More precisely, PIMT levels were significantly lower by 76% in pilocytic astrocytomas (grade I), 46% in astrocytomas (grade II), 69% in anaplastic astrocytomas (grade III), and a marked 80% in glioblastomas (grade IV) as compared to normal brains. RT-PCR analysis showed that levels of type I PIMT mRNA were up-regulated while those of type II PIMT mRNA were down-regulated in glioblastomas. Furthermore, the reduced PIMT levels correlated closely with a decrease in the number of neuron cells in astrocytic tumors as assessed by measuring the neuron-specific enolase level. Many proteins with abnormal aspartyl residues accumulated in brain tumors and some were specific to individual grades of astrocytic tumors. Similar results were obtained, either by measuring the reduction in PIMT activity and expression or by measuring the formation of abnormal proteins, in an orthotopic rat brain tumor model implanted with invasive CNS-1 glioma cells. The novelty of these findings was to provide the first evidence for a marked reduction of PIMT expression and activity during stage progression of astrocytic tumors in humans.

Regulatory mechanisms controlling biogenesis of ubiquitin and the proteasome

Analysis of several Saccharomyces cerevisiae ump mutants with defects in ubiquitin (Ub)-mediated proteolysis yielded insights into the regulation of the polyubiquitin gene UBI4 and of proteasome genes. High-molecular weight Ub–protein conjugates accumulated in ump mutants with impaired proteasome function with a concomitant decrease in the amount of free Ub. In these mutants, transcriptional induction of UBI4 was depending in part on the transcription factor Rpn4. Deletion of UBI4 partially suppressed the growth defects of ump1 mutants, indicating that accumulation of polyubiquitylated proteins is deleterious to cell growth. Transcription of proteasome subunit genes was induced in ump mutants affecting the proteasome, as well as under conditions that mediate DNA damage or the formation of abnormal proteins. This induction required the transcriptional activator Rpn4. Elevated Rpn4 levels in proteasome-deficient mutants or as a response to abnormal proteins were due to increased metabolic stability. Up-regulation of proteasome genes in response to DNA damage, in contrast, is shown to operate via induction of RPN4 transcription.

Aggresome Formation in Liver Cells in Response to Different Toxic Mechanisms: Role of the Ubiquitin-Proteasome Pathway and the Frameshift Mutant of Ubiquitin

Aggresomes form in cells when intracellular proteins undergo conformational changes, as in so-called conformational diseases. This phenomenon has been observed in the liver and brain and in cell culture in response to abnormal protein formation, such as mutant proteins. In the case of the brain the frameshift mutant ubiquitin (UBB+1) is involved. Mallory body formation in the liver is one example of this phenomenon in vivo. Mallory body formation is common in a variety of liver diseases of diverse pathogenesis. The study of the Mallory body forming model indicated that drug-conditioned hepatocytes form Mallory bodies when mice are given colchicine, ethanol, okadaic acid, or exposure to heat shock. These findings suggest that aggresome formation is a common pathway of liver injury due to diverse mechanisms. To further characterize the role of this common pathway, drug-primed mice were exposed to different types of liver injury, i.e., using such drugs as thioacetamide, galactosamine, tautomycin, and the proteasome inhibitor PS341. Mallory body formation was induced by treatment with all the toxins tested, giving credence to the proposal that aggresome formation in the liver is a common pathway in response to different primary mechanisms of liver injury. The frameshift mutant UBB+1 was invariably found to colocalize with ubiquitin in the Mallory body, indicating its essential involvement in the mechanism of MB formation.

Expression of heat shock proteins in Alzheimer's disease

In an investigation of heat shock proteins (HSPs) in the brains of Alzheimer's disease (AD) patients and cognitively intact control subjects, we found that 2 HSPs, termed "HSP72" and "GRP78," underwent major changes in expression in AD. HSP72, which was present at very low levels in control brains, increased dramatically in AD patients, and was localized exclusively in neuritic plaques and neurofibrillary tangles. We hypothesize that HSP72 is induced as an early response to the formation of abnormal proteins, perhaps targeting them for proteolysis. In contrast, GRP78 increased in AD only in neurons that remained cytologically normal, especially in the CA3 subfield of the hippocampus and the deep layers of the entorhinal cortex. The increased expression of GRP78 within successfully surviving neurons suggests that this protein may protect such cells from AD-specific damage.

Age-dependent effect of UV light in abnormal alpha neoprotein formation in the lens

The aim of this study was to determine whether or not alpha neoprotein formation can occur in the lens in response to a photolytic process. Alpha crystallin from young calf and aged bovine lenses was irradiated for up to 22 hours with UV light. After cortical alpha crystallin had been irradiated, it completely preserved its quaternary antigenic determinants. In contrast, 24.3% of the bovine nuclear alpha crystallin completely lost their quaternary determinants as a result of irradiation. The latter population of molecules still preserved the association of A chains with B chains, suggesting the formation of abnormal proteins. The exposure to UV irradiation can cause the formation of alpha neoproteins from aged but not from young alpha crystallin. Comparison of photolytic rates based on spectroscopic and chromatographic parameters also show an age-related increase in the susceptibility of alpha crystallin to photolyze. It is suggested that this increase is due to the age-related formation of a photosensitizer in alpha crystallin.

Etiologic theories of Alzheimer's disease

A variety of theories about the cause or origin of Alzheimer's disease (AD) have been proposed. Some of these theories are based on a particular research finding, such as abnormal protein formation or aluminum accumulation. Others are based on possible imbalances in brain function (calcium disruption, for example), and some are based on analogies to other diseases such as infectious disorders. Still other theories are based on general systemic conditions such as immune function and nutrition, and some are simply based on speculation. Evidence has been found for separate classifications or subgroups of AD that could imply separate causes. There have also been various genetic explanations for the familial category of AD. This review summarizes the various theories suggested for AD with the reason sfor their proposal, the evidence for or against them, and their current status. Appraisals have been carnied out from the perspective that identification of the primary cause of AD could be crucial to the discovery of effective treatment strategies.

Immunochemical analysis of the turnover of ubiquitin-protein conjugates in intact cells. Relationship to the breakdown of abnormal proteins.

Previous studies in a cell-free proteolytic system from reticulocytes indicated that the conjugation of ubiquitin with proteins plays a role in protein breakdown. To examine some of the physiological functions of the ubiquitin conjugation system, and immunochemical method was developed for the isolation of ubiquitin-protein conjugates from intact cells. A specific antiserum was raised against ubiquitin and purified by affinity chromatography on ubiquitin-Sepharose. When cells are labeled with tryptophan (which is missing from ubiquitin), labeled immunoreactive material isolated by the antibody is derived from the protein moiety of ubiquitin-protein conjugates. There is a marked increase in the labeling of ubiquitin-protein conjugates during the formation of abnormal proteins in reticulocytes (induced by the incorporation of amino acid analogs), suggesting that proteins with abnormal structure are more readily conjugated to ubiquitin than most normal proteins. Essentially similar, although less marked, effects of amino acid analogs were observed in Ehrlich ascites cells. When further protein synthesis was blocked with cycloheximide, ubiquitin conjugates decayed more extensively than the corresponding average labeled cellular proteins. This is consistent with the interpretation that a considerable part of ubiquitin conjugates is derived from a pool of rapidly degradable proteins.

The biochemical pathology of ethionine-lnduced pancreatic damage: Ethionine incorporation into proteins, and ATP levels in pancreas of rats

Two biochemical mechanisms of possible pathogenetic significance in injury of cells by ethionine are synthesis of abnormal proteins by ethionine incorporation, and induction of ATP deficiency. These hypotheses were evaluated in relation to ethionine-in-duced pancreatic damage in rats. Ethionine incorporation into pancreatic protein was found to conform to the pattern which has been reported for several natural amino acids. Incorporation continued for at least 9 hours following a dose of 0.5 g per kg, apparently because a significant level of free ethionine persisted. The pancreas did not appear to have any unique tendency to incorporate ethionine into structural proteins when compared with liver and kidney, and therefore the results do not suggest that formation of abnormal proteins is of special significance in the pancreas. S-adenosylethionine accumulation and ATP deficit were demonstrated in pancreas 24 hours following single doses of ethionine (1 g per kg). However, at a lower dose which was adequate to produce severe pancreatic damage, there was no decrease in ATP in the pancreas. It therefore appears that the ethionine-induced pancreatic lesion cannot be attributed to ATP deficiency.

Analog-induced inclusions in pancreatic acinar cells

Feeding of β-3-thienyl-dl-alanine in semisynthetic diets devoid of phenylalanine produces characteristic focal areas of cytoplasmic necrosis. The cytoplasmic components within sequestered areas give rise to myeloid structures and small granules within complex dense bodies. Further alteration of the contents of complex dense bodies results in appearance of homogeneous bodies. This process appears to be related to a disturbed protein synthesis and to formation of abnormal proteins; it may represent a mechanism by which the cell eliminates abnormal proteins of endogenous and exogenous origin.

Nature of increased protein and DNA in chronic hepatic injury.

EXCESS protein and deoxyribonucleic acid (DNA) are found in chronic hepatic injury, for example, after administration of ethionine1–3, butter yellow4,5, 2 acetyl aminofluorene6,7, and a high fat–low choline–low protein diet1. This suggested that the excess protein or DNA is an abnormal product of the hepatocytes1–3. This is supported by the incorporation of ethionine into hepatic, proteins2, presumably the abnormal proteins. These proteins, or DNA, have been related to cancer1,7. This communication is to point out that the excess protein formation results from alterations in cell population, rather than from formation of abnormal proteins.

Influence of hepatectomy and hepatic necrosis on electrophoresis of plasma proteins.

With the advent of physicochemical methods of preparative chromatography and electrophoresis, considerable information relating to plasma protein patterns in normal and disease states has evolved. 12 Although it appears that the liver is the source of all the plasma albumin, fibrinogen, and most of the globulin, 6 its role in the aberration of these protein patterns in disease as shown by electrophoresis has not been clearly defined. It remains unclear whether abnormal patterns are due to lack of formation of proteins by the liver, to the formation of abnormal proteins either by the liver or by the extrahepatic tissues, or to the conjugation of normal proteins with abnormal metabolites, thereby altering their physical properties. To aid in answering these questions, the changes in the electrophoretic patterns of the plasma proteins were studied for varying periods following surgical removal of all or part of the liver and after chemical hepatic necrosis