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At least 19 recordsLinked to original sources
PubMed2026

Mapping self-associating chromatin hubs identifies Id proteins as key determinants of exhausted CD8+ T cell fate.

Within days of exposure to chronic viral infections, activated CD8+ T cells differentiate into Tcf1-Slamf6loTim3hi exhaustion-prone effector T (TEX_EFF) cells or self-renewing Tcf1+Slamf6hiTim3lo precursor exhausted T (TPEX) cells. Here we showed that early CD8+ TEX cell fates were imprinted by forming subset-specific, self-associating chromatin hubs. Chromatin hub assembly coincided with effector or stemness gene induction and identified the transcription cofactors Id2 and Id3 as key regulators that promoted CD8+ TEX_EFF and CD8+ TPEX cell fates, respectively. Id2 drove CD8+ TEX_EFF cell specification by activating effector genes, while suppressing genes involved in exhaustion and stemness. In contrast, Id3-repressed effector genes but upregulated IL-7Rα and AhR, thereby maintaining the CD8+ TPEX cell pool. Mechanistically, Id2 and Id3 exhibited a distinct impact on the chromatin accessibility landscape in early CD8+ TEX cells by engaging Runx3 and Tcf1 transcription factors along with E proteins. These findings indicated that reshaping chromatin architecture represents a critical means for specifying CD8+ TEX cell fates and ensuring lineage stability.

Animals↗
PubMed2025

Guanine nucleotide biosynthesis blockade impairs MLL complex formation and sensitizes leukemias to menin inhibition.

Targeting the dependency of MLL-rearranged (MLLr) leukemias on menin with small molecule inhibitors has opened new therapeutic strategies for these poor-prognosis diseases. However, the rapid development of menin inhibitor resistance calls for combinatory strategies to improve responses and prevent resistance. Here we show that leukemia stem cells (LSCs) of MLLr acute myeloid leukemia (AML) exhibit enhanced guanine nucleotide biosynthesis, the inhibition of which leads to myeloid differentiation and sensitization to menin inhibitors. Mechanistically, targeting inosine monophosphate dehydrogenase 2 (IMPDH2) reduces guanine nucleotides and rRNA transcription, leading to reduced protein expression of LEDGF and menin. Consequently, the formation and chromatin binding of the MLL-fusion complex is impaired, reducing the expression of MLL target genes. Inhibition of guanine nucleotide biosynthesis or rRNA transcription further suppresses MLLr AML when combined with a menin inhibitor. Our findings underscore the requirement of guanine nucleotide biosynthesis in maintaining the function of the LEDGF/menin/MLL-fusion complex and provide a rationale to target guanine nucleotide biosynthesis to sensitize MLLr leukemias to menin inhibitors.

Proto-Oncogene Proteins↗
PubMed1979

The identification of distinctive forms of human alpha 2-macroglobulin by using the numerical relationship between trypsin binding in alpha- and beta-modes.

Interactions between the serine proteinase trypsin and the protein proteinase inhibitors in human blood were expressed in terms of a coupled set of non-linear differential equations, which has been solved for each of 110 samples of serum obtained from colleagues and from a variety of hospital sources. Optimization of nine unknown theoretical parameters and 21 experimental rate measurements of the hydrolytic activity of trypsin in free and bound states after admixture with various amounts of a given serum was achieved by an iterative procedure using initial estimates of the parameters derived from the "four-straight-line" model described in the preceding paper [Topping & Seilman (1979) Biochem. J. 177, 493--499.] Such a procedure yielded the following information for each sample of serum examined: (a) the concentrations of alpha 1-antitrypsin and alpha 2-macroglobulin; (b) the unequivocal assignment of alpha 2-macroglobulin into one of seven categories on the basis of trypsin binding in two kinetically differentiated modes (alpha and beta); (c) the hydrolytic activities of trypsin (versus Bz-Arg-OEt) when bound to alpha 1-antitrypsin, and to alpha 2-macroglobulin in the alpha- and beta-modes. Molecular interpretations of the binding of trypsin to alpha 2-macroglobulin are discussed and the potential clinical value of recognizing the nature of such binding is reported.

Animals↗
PubMed2026

Proteomic Immune Signatures of Severe HIV-Associated Tuberculosis in Sub-Saharan Africa: A Prospective, Multicenter Analysis From Uganda.

OBJECTIVES: Severe tuberculosis (TB) is a major cause of critical illness and death in people living with HIV (PLWH) worldwide. Despite this, the immunopathology of severe HIV-associated TB (HIV/TB) is poorly understood. We aimed to identify an immunopathologic signature of severe HIV/TB in sub-Saharan Africa. DESIGN AND SETTING: We analyzed proteomic data from two prospective observational cohorts of adults hospitalized with severe undifferentiated infection in Uganda: an urban discovery cohort (Entebbe, n = 241) and a rural validation cohort (Tororo, n = 253). PATIENTS: Adults (age ≥ 18 yr) hospitalized with severe febrile illness. INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: Across both cohorts, severe HIV/TB was common, affecting 18% of participants in the discovery cohort and 21% in the validation cohort. Overall mortality was significant (30-d mortality of 22% in the discovery cohort and 60-d mortality of 26% in the validation cohort). Participants were stratified into three HIV/TB phenotypes: HIV-negative without TB, PLWH without TB, and PLWH with microbiologically diagnosed TB. We applied ordinal random forest models in the discovery cohort as a supervised feature-selection approach to identify proteins associated with progressive HIV/TB phenotype. In both cohorts, PLWH with microbiologically diagnosed TB were at highest risk of critical illness and death (30-d mortality of 42% in the discovery cohort and 60-d mortality of 52% in the validation cohort). An eight-protein signature reliably distinguished this phenotype, reflecting mediators of macrophage/dendritic cell activation (lysosome-associated membrane glycoprotein 3), natural killer cell and T-cell stimulation and cytotoxicity (cluster of differentiation 70, class I-restricted T-cell-associated molecule), B-cell activation (immunoglobulin lambda constant 2), protease-mediated tissue injury (protease, serine 2 [trypsin-2]), dysregulated coagulation (serpin peptidase inhibitor, clade A [alpha-1 antitrypsin], member 5), extracellular matrix remodeling (epidermal growth factor-containing fibulin-like extracellular matrix protein 1), and growth hormone/insulin-like growth factor axis dysregulation (insulin-like growth factor binding protein 3). CONCLUSIONS: We identified an immunologic signature of severe HIV/TB defined by mediators of macrophage/dendritic cell and cytotoxic lymphocyte activation, extracellular matrix remodeling, and dysregulated coagulation. These findings offer new insight into HIV/TB pathobiology and highlight potential targets for host-directed therapies in this high-risk population.

Humans↗
PubMed2025

Reprogramming neuroblastoma by diet-enhanced polyamine depletion.

Neuroblastoma is a highly lethal childhood tumour derived from differentiation-arrested neural crest cells1,2. Like all cancers, its growth is fuelled by metabolites obtained from either circulation or local biosynthesis3,4. Neuroblastomas depend on local polyamine biosynthesis, and the inhibitor difluoromethylornithine has shown clinical activity5. Here we show that such inhibition can be augmented by dietary restriction of upstream amino acid substrates, leading to disruption of oncogenic protein translation, tumour differentiation and profound survival gains in the Th-MYCN mouse model. Specifically, an arginine- and proline-free diet decreases the amount of the polyamine precursor ornithine and enhances tumour polyamine depletion by difluoromethylornithine. This polyamine depletion causes ribosome stalling, unexpectedly specifically at codons with adenosine in the third position. Such codons are selectively enriched in cell cycle genes and low in neuronal differentiation genes. Thus, impaired translation of these codons, induced by combined dietary and pharmacological intervention, favours a pro-differentiation proteome. These results suggest that the genes of specific cellular programmes have evolved hallmark codon usage preferences that enable coherent translational rewiring in response to metabolic stresses, and that this process can be targeted to activate differentiation of paediatric cancers.

Animals↗
PubMed1976

A labile acid phosphatase isozyme associated with the surface of vegetative Dictyostelium discoideum cells.

A minor acid phosphatase isozyme (acid phosphatase I) of vegetative Dictyostelium discoideum amebae has been shown to be associated exclusively with the external surface of the plasma membrane. The isozyme is not present in phagocytic vacuoles isolated with latex beads. The isozyme disappears from cells removed from nutrient medium and does not reappear during differentiation. When inhibitors of protein synthesis (e.g. cycloheximide, chloral hydrate, concanavalin A) are added to cells growing in nutrient medium, acid phosphatase I is rapidly lost. It appears that the level of protein synthesis need only be moderately reduced (less than 25%) to induce loss of enzyme activity. Treatment with inhibitors of DNA and RNA synthesis for up to 2 h had no effect on isozyme activity. It is postulated that the cells are able to "sense" (through the reduction in levels of protein synthesis) when external conditions become unfavorable, and immediately respond by reducing the activity of enzymes involved in maintaining contact with the extracellular environment. The closed system thought necessary for differentiation would then be created.

Acid Phosphatase↗
PubMed1979

Differential effects of two interferon-induced translational inhibitors on initiation of protein synthesis.

At least two different mechanisms for the inhibition of mRNA translation operate in extracts of interferon-treated L cells. One is mediated by an interferon-induced protein kinase which, when activated by double-stranded RNA and ATP, phosphorylates the small subunit of initiation factor eIF-2. Addition of the purified interferon-induced protein kinase to L cell extracts, strongly reduces the amount of methionyl-tRNA bound to 40-S ribosomal subunits. The second translational inhibition is due to the synthesis of (2'-5')oligo(adenylate) by interferon-induced enzyme E. The oligonucleotide in turn activates a ribonuclease F constitutively present in L cells. Addition of the purified nuclease with its oligonucleotide activator to L cell extracts produces a strong decrease in polyribosome formation and an accumulation of initiation complex. These experiments differentiate the effects of the two interferon-induced inhibitors on mRNA translation.

Animals↗
PubMed1976

Regulation of immunity and inflammation by mediators from macrophages.

Mononuclear phagocytes secrete a number of materials into the extracellular environment. The materials secreted by phagocytes can be grouped into three categories: a) enzymes affecting extracellular proteins (collagenase, elastase, lysosomal proteases, plasminogen activators), b) materials involved in defense processes (complement proteins, interferons, lysozyme), and c) factors regulating activities of surrounding cells. The latter include lymphostimulatory molecules, a colony-stimulating factor, and inhibitors of cell growth. The conditions for secretion of the materials depend on the activity of the phagocytes. The lymphostimulatory molecules secreted by macrophages exert various effects: 1) an increase in DNA synthesis of lymphocytes, 2) a maturation of early thymocytes to mature T cells, and 3) the differentiation of some B cells to antibody-secreting cells. The mitogenic principle has been partially isolated as a protein of 15,000 to 20,000 daltons. The secretion of lymphostimulatory molecules is increased following uptake of various materials by macrophages or by addition of activated T cells to macrophage cultures.

Animals↗
PubMed1976

Different susceptibility of protein synthesis to inhibitors of elongation in cell-free systems from plasma cell tumours and reticulocytes.

Plasma cells and reticulocytes are mammalian cell systems which have specialized in the synthesis of a single protein during their differentiation from one common stem cell. To study whether there is a difference in cell susceptibility at the level of elongation, dose vs. inhibition curves of sparsomycin, cycloheximide and emetine in cell-free systems with S-30 fractions from plasma cell tumours (MOPC 63, MOPC 41, RPC 20, MOPC 104 E), reticulocytes and liver were compared. The experiments revealed: (1) all the selected systems are equally sensitive to sparsomycin; (2) the susceptibility of the reticulocyte systems to cycloheximide and emetine is higher than that of the plasma cell tumours. In the dose range of 1 - 10(-7) --5 - 10(-5) M cycloheximide and 1 - 10(-6)--1 - 10(-4) M emetine the reticulocyte system is preferentially inhibited; (3) the sensitivities of all plasma cell tumours are equal; (4) the liver system is more sensitive to emetine than to cycloheximide; (5) the site of the different susceptibility to these antibiotics could be located on the ribosomes; (6) however, when the extracts of the plasma cell tumours were prepared in the presence of hemin, their susceptibility rises and is like that of reticulocytes. These results show that hemin promotes in a cell-specific manner the sensitivity to some inhibitors of protein synthesis.

Animals↗
PubMed1978

Association of Tenebrio molitor L. alpha-amylase with two protein inhibitors--one monomeric, one dimeric--from wheat flour. Differential scanning calorimetric comparison of heat stabilities.

Thermal stabilization resulting from protein . protein association between two protein inhibitors (coded as 0.19, a dimer, and 0.28, a monomer) from wheat flour and the alpha-amylase from Tenebrio molitor L. (yellow mealworm) larvae was investigated by differential scanning calorimetry (heating rate 10 degrees C/min). Thermograms (plots of heat flow vs. temperature) for the two inhibitors showed broad endothermic peaks with the same extrema (denaturation temperatures) at 93 degrees C, and equal, small enthalpies of denaturation (2 cal/g). The amylase produced a sharp endotherm at 70.5 degrees C, but a larger enthalpy change on denaturation (6 cal/g). The amylase . inhibitor complexes differed in thermal stability, but both showed significant stabilization relative to free enzyme. The complex formed with monomeric inhibitor 0.28 showed a higher denaturation temperature (85.0 degrees C) than that formed with dimeric inhibitor 0.19 (80.5 degrees C). This order of stabilization agrees with the relative affinities of the inhibitors for the amylase. These thermograms are consistent with previous results which indicated that 1 mol of amylase binds 1 mol of inhibitor 0.19.

Albumins↗
PubMed2025

Hypoxia-inducible factor 2 regulates alveolar regeneration after repetitive injury in three-dimensional cellular and in vivo models.

Idiopathic pulmonary fibrosis (IPF) is a progressive interstitial lung disease in which repetitive epithelial injury and incomplete alveolar repair result in accumulation of profibrotic intermediate/transitional "aberrant" epithelial cell states. The mechanisms leading to the emergence and persistence of aberrant epithelial populations in the distal lung remain incompletely understood. By interrogating single-cell RNA sequencing (scRNA-seq) data from patients with IPF and a mouse model of repeated lung epithelial injury, we identified persistent activation of hypoxia-inducible factor (HIF) signaling in these aberrant epithelial cells. Using mouse genetic lineage-tracing strategies together with scRNA-seq, we found that these disease-emergent aberrant epithelial cells predominantly arose from airway-derived (Scgb1a1-CreER-traced) progenitors and exhibited transcriptional programs of Hif2a activation. In mice treated with repetitive intratracheal bleomycin, deletion of Epas1 (Hif2a) but not Hif1a, from airway-derived progenitors, or administration of the small-molecule HIF2 inhibitor PT-2385, using both prevention and rescue approaches, attenuated experimental lung fibrosis, reduced the appearance of aberrant epithelial cells, and promoted alveolar repair. In mouse alveolar organoids, genetic or pharmacologic inhibition of Hif2 promoted alveolar differentiation of airway-derived epithelial progenitors. In addition, treatment of human distal lung organoids with PT-2385 increased colony-forming efficiency, enhanced protein and transcriptional markers of alveolar type 2 epithelial cell maturation, and prevented the emergence of aberrant epithelial cells. Together, these studies showed that HIF2 activation drives the emergence of aberrant epithelial populations after repetitive injury and that targeted HIF2 inhibition may represent an effective therapeutic strategy to promote functional alveolar repair in IPF and other interstitial lung diseases.

Animals↗
PubMed1979

Characterisation of protein synthesis in cell-free extracts from different mammalian cells by their sensitivity to inhibitors of polypeptide-chain initiation.

Plasma cells and reticulocytes are differentiated mammalian cell systems specialized in the synthesis of distinct proteins. To study whether a cell specificity of polypeptide-chain initiation in such cell systems can be detected through inhibitors, the sensitivity to several drugs interfering with initiation was compared in cell-free systems with S-30 extracts from these cells. The following was indicated by the experiments: (1) under the selected conditions, the different cell-free systems are comparable with respect to their activity of initiation, and that aurintricarboxylic acid inhibits mRNA binding to the ribosome in plasma cell tumours the same as in reticulocytes. (2) The sensitivity of translation of endogenous mRNAs in reticulocytes and plasma cell tumours to inhibitors of mRNA binding to the ribosome, aurintricarboxylic acid and the homopolynucleotides poly(U) and poly(A), are different. (3) Inhibition of the succeeding reactions of peptide-chain initiation by sodium fluoride and pactamycin was not selective. (4) In both cell systems translation of poly(U) is equally sensitive to aurintricarboxylic acid. (5) In extracts from TEPC 15 myeloma cells synthesis of immunoglobulin L-chains compared with that of the other myeloma proteins is partially resistant to aurintricarboxylic acid, whereas in reticulocytes, no differential sensitivities of individual proteins could be observed. The different susceptibility of the mRNA binding reaction in plasma cell tumours and reticulocytes suggests that the predominant mRNAs of these cells have different affinities to ribosome binding sites.

Animals↗
PubMed1979

Gametocytogenesis of Plasmodium falciparum in vitro: the cell-cycle.

Reproducible growth of gametocytes of Plasmodium falciparum in vitro was obtained from ring-stages taken directly from naturally infected patients and from the same material following storage in liquid nitrogen. Progressive sexual differentiation in vitro was examined for a finite period of 9 days in microcultures and was, for convenience, divided into 5 stages using established morphological criteria (Hawking, Wilson & Gammage, 1971). This microculture system was adapted as a bioassay for various anti-metabolites. Drug activity was measured by observing the inhibition of the established pattern of sequential development in experimental as compared to control cultures. Inhibitors used were directed against DNA, RNA and protein metabolism and microtubule assembly. As a result of these studies it is proposed that the sexual cell-cycle of P. falciparum is characterized by 4 phases. (1) A G1 period which lasts only a few hours. (2) The S phase, where DNA synthesis occurs, occupies the remainder of the first 2 days of development - both G1 and S are confined to stage I and II gametocytes. (3) G2, which is subdivided into 2 sections: G2A, characterized by stage II and III gametocytes, in which significant RNA and protein synthesis continue to occur; and G2B, where there is a progressive increase in transcription control resulting in the depression of both RNA and protein synthesis. Nonetheless, continued morphological differentiation occurs in the latter section transforming the parasites to stage IV and the morphologically and functionally mature stage V. The final M phase is marked by the brief and exposive events of gametogenesis, during which further protein synthesis occurs de novo. The proposed cell-cycle is examined as a model for studies on the activity of gametocytocidal compounds.

Animals↗
PubMed1978

Enumeration and ultrastructure of C4-producing free alveolar cells from guinea pig lung.

Free alveolar cells from guinea pig lung producing the fourth emoponent of C (C4) were identified, enumerated, and characterized by using anti-C4 Fab-peroxidase conjugates in conjunction with transmission electron microscopy. The C4-producing cell population consisted of: 1) alveolar macrophages (AM); 2) less well differentiated phagocytes similar in morphology to exudate macrophages; and 3) weakly phagocytic secretory cells with numerous profiles of rough-surfaced endoplasmic reticulum (ER). Internal immunolabeling allowed the visualization of C4 in the ER, perinuclear space, and Golgi complex of producer cells and its release at cell surfaces; synthesis of C4 in vitro was sensitive to inhibitors both of protein synthesis and messenger RNA function. The percentage of free alveolar cells from normal animals competent for C4 production as indicated by cell surface immunolabeling was approximately 1% of the total cells obtained by lavage. Transnasal infection with Listeria monocytogenes, generation of a pulmonary granulomatous reaction by i.v. injection of heat-killed BCG, and aerosol infection of nonvaccinated animals with Myco-bacterium tuberculsois each resulted in an increase in numbers of AM and exudate macrophage-like free alveolar cells competent for C4-production.

Animals↗
PubMed1975

Differentiation between binding and transport of dansylgalactosides in Escherichia coli.

The results presented in this paper confirm and extend previous observations which indicate that fluorescent dansylgalactsodes bind to the beta-galactoside carrier protein but do not penetrate the cytoplasmic membrane. The conclusion is supported by the following observations. (a) Although 2'-(N-dansyl)aminoethyl-beta-D-thiogalactopyranoside and 2'-(N-dansyl)aminoethyl-beta-D-galactopyranoside are competitive inhibitors of lactose transport in intact cells of Escherichia coli and induce the in vitro synthesis of beta-galactosidase, they do not induce beta-galactosidase in vivo. (b) p-Chloromercuribenzenesulfonate does not cause efflux of lactose from the intravesicular pool, but causes rapid reversal of D-lactate-induced dansylgalactoside fluorescence. (c) Dansylgalactosides inhibit dilution-induced, carrier-mediated lactose efflux.

4-Chloromercuribenzenesulfonate↗
PubMed1979

Regulation of dome formation in differentiated epithelial cell cultures.

Rat mammary (Rama 25) and dog kidney (MDCK) epithelial cell cultures formed 'domes' of cells due to fluid accumulation in focal regions between the culture dish and the cell monolayer. Addition of ouabain caused collapse of domes, suggesting that transport functions were required for maintenance of domes. Dome formation in both epithelial cell lines was stimulated by a broad spectrum of known inducers of erythroid differentiation in Fried erythroleukemia cells. Among these inducers were: 1) polar solvents such as dimethyl-sulfoxide, dimethylformamide, and hexamethylene bisacetamide; 2) purines such as hypoxanthine, inosine, and adenosine; 3) low-molecular-weight fatty acids such as n-butyrate; and 4) conditions expected to elevate levels of cyclic AMP. In the latter group were activators of adenylate cyclase such as cholera toxin and prostaglandin E 1; cyclic AMP phosphodiesterase inhibitors such as theophylline and 1-methyl-3-isobutylxanthine; and analogs of cyclic AMP. Induction of domes occurred 15--30 h after addition of inducer to the culture medium. Induction by chemicals was serum-dependent and required protein synthesis but not DNA synthesis. Induced dome formation was reversible after removal of inducer, requiring the continuous presence of inducer. Reversal was also observed after either either removal of serum or addition of inhibitors of protein synthesis. These results suggest that hypothesis that domes arise in these epithelial cultures by a process that is similar to cell differentiation and is influenced by cyclic AMP.

Animals↗
PubMed1977

Specificity of a protein phosphatase inhibitor from rabbit skeletal muscle.

A hear-stable protein, which is a specific inhibitor of protein phosphatase-III, was purified 700-fold from skeletal muscle by a procedure that involved heat-treatment at 95 degrees C, chromatography on DEAE-cellulose and gel filtration on Sephadex G-100. The final step completely resolved the protein phosphatase inhibitor from the protein inhibitor of cyclic AMP-dependent protein kinase. The phosphorylase phosphatase, beta-phosphorylase kinase phosphatase, glycogen synthase phosphatase-1 and glycogen synthase phosphatase-2 activities of protein phosphatase-III [Antoniw, J. F., Nimmo, H. G., Yeaman, S. J. & Cohen, P.(1977) Biochem.J. 162, 423-433] were inhibited in a very similar manner by the protein phosphatase inhibitor and at least 95% inhibition was observed at high concentrations of inhibitor. The two forms of protein phosphatase-III, termed IIIA and IIIB, were equally susceptible to the protein phosphatase inhibitor. The protein phosphatase inhibitor was at least 200 times less effective in inhibiting the activity of protein phosphatase-I and protein phosphatase-II. The high degree of specificity of the inhibitor for protein phosphatase-III was used to show that 90% of the phosphorylase phosphatase and glycogen synthase phosphatase activities measured in muscle extracts are catalysed by protein phosphatase-III. Protein phosphatase-III was tightly associated with the protein-glycogen complex that can be isolated from skeletal muscle, whereas the protein phosphatase inhibitor and protein phosphatase-II were not. The results provide further evidence that the enzyme that catalyses the dephosphorylation of the alpha-subunit of phosphorylase kinase (protein phosphatase-II) and the enzyme that catalyses the dephosphorylation of the beta-subunit of phosphorylase kinase (protein phosphatase-III) are distinct. The results suggest that the protein phosphatase inhibitor may be a useful probe for differentiating different classes of protein phosphatases in mammalian cells.

Animals↗
PubMed1977

Identification of "angiotensin immunoreactive material" in rat kidney.

The reported presence of large quantities of a high molecular weight form of angiotensin I and II in renin granules from rat kidney cortex was investigated. Subcellular fractionation by differential centrifugation and isopycnic gradient centrifugation confirmed the presence of 'angiotensin I immunoreactive material,' but the distribution resembled that of lysosomes rather than renin granules, mitochondria or protein. The material did not possess pressor properties, was stable to incubation at 37 C and was precipitated by ethanol. Incubation of subcellular fractions with peptidase inhibitors known to inhibit the activity of renal angiotensinases (EDTA, diisopropyl phosphorofluoridate and 2,3-dimercaptopropanol) decreased the level of 'angiotensin I immunoreactive material' in kidney fractions treated by radioimmunoassay. By paper chromatography it was apparent that subcellular fractions were capable of degrading 125I-labelled angiotensin I used as tracer in the radioimmunoassay. The degree of degradation followed the distribution of lysosomes among the fractions and was decreased by the angiotensinase inhibitors. The apparent molecular weight of the major portion of angiotensinase activity persisting despite the presence of angiotensinase inhibitors was 75,000, a value similar to that of 'angiotensin immunoreactive material'inkidney fractions treated with non-ionic detergent. On the basis of the present findings it is suggested that 'angiotensin immunoreactive material' may be largely artifactual, beingcreated by the hydrolysis of tracer by angiotensinase enzymes during radioimmunoassay to form fragments that are no longer capable of binding to the specific antibody. This would produced results that would appear the same as those produced had genuine angiotensin immunoreactivity indeed been present in the samples. Such an effect could, in principle, occur in any competitive protein binding assay and should be tested.

Angiotensin II↗