scholarly journals Ganglioside GM2-Activator Protein and Vesicular Transport in Collecting Duct Intercalated Cells

1999 ◽  
Vol 10 (3) ◽  
pp. 435-443
Author(s):  
THOMAS M. MUNDEL ◽  
HANS W. HEID ◽  
DON J. MAHURAN ◽  
WILHELM KRIZ ◽  
PETER MUNDEL
Keyword(s):  
Collecting Duct ◽  
Rat Kidney ◽  
The Body ◽  
Kidney Cortex ◽  
Intercalated Cells ◽  
Rat Kidney Cortex ◽  
Activator Protein ◽  
Hexosaminidase A ◽  
Gm2 Activator ◽  
Gm2 Activator Protein

Abstract. This study describes the molecular characterization of an antigen defined by an autoantibody from a woman with habitual abortion as GM2-activator protein. The patient showed no disorder of renal function. Accidentally with routine serum screening for autoantibodies, an immunoreactivity was found in kidney collecting duct intercalated cells. Three distinct patterns of immunostaining of intercalated cells were observed: staining of the apical pole, basolateral pole, and diffuse cytoplasmic labeling. Ultrastructurally, the immunoreactivity was associated with “studs,” which represent the cytoplasmic domain of the vacuolar proton pump in intercalated cells. This pump is subjected to a shuttling mechanism from cytoplasmic stores to the cell membrane, which exclusively occurs in intercalated cells. Peptide sequences of a 23-kD protein purified from rat kidney cortex showed complete identity with corresponding sequences of GM2-activator protein. In the brain, GM2-activator protein is required for hexosaminidase A to split a sugar from ganglioside GM2. Because neither ganglioside GM2 nor GM1 (its precursor) is present in significant amounts in the kidney, the previous finding that this tissue contains the highest level of activator protein in the body was confusing. In this study, a novel role for GM2-activator protein in intercalated cells is proposed, and possible roles in the shuttling mechanism are discussed.

scholarly journals In Cellulo Examination of a Beta-Alpha Hybrid Construct of Beta-Hexosaminidase A Subunits, Reported to Interact with the GM2 Activator Protein and Hydrolyze GM2 Ganglioside

2021 ◽  
Author(s):  
Warren W. Wakarchuk ◽  
Incilay Sinici ◽  
Sayuri Yonekawa ◽  
Ilona Tkachyova ◽  
Steven J. Gray ◽  
...  
Keyword(s):  
In Vitro Assays ◽  
Variant Form ◽  
Gm2 Ganglioside ◽  
Activator Protein ◽  
Hexosaminidase A ◽  
Hybrid Construct ◽  
Gm2 Activator ◽  
Gm2 Activator Protein

The hydrolysis in lysosomes of GM2 ganglioside to GM3 ganglioside requires the correct synthesis, intracellular assembly and transport of three separate gene products; i.e., the alpha and beta subunits of heterodimeric beta-hexosaminidase A, E.C. # 3.2.1.52 (encoded by the HEXA and HEXB genes, respectively), and the GM2-activator protein (GM2AP, encoded by the GM2A gene). Mutations in any one of these genes can result in one of three neurodegenerative diseases collectively known as GM2 gangliosidosis (HEXA, Tay-Sachs disease, MIM # 272800; HEXB, Sandhoff disease, MIM # 268800; and GM2A, AB-variant form, MIM # 272750). Elements of both of the hexosaminidase A subunits are needed to productively interact with the GM2 ganglioside-GM2AP complex in the lysosome. Some of these elements have been predicted from the crystal structures of hexosaminidase and the activator. Recently a hybrid of the two subunits has been constructed and reported to be capable of forming homodimers that can perform this reaction in vivo, which could greatly simplify vector-mediated gene transfer approaches for Tay-Sachs or Sandhoff diseases. A cDNA encoding a hybrid hexosaminidase subunit capable of dimerizing and hydrolyzing GM2 ganglioside could be incorporated into a single vector, whereas packaging both subunits of hexosaminidase A into vectors, such as adeno-associated virus, would be impractical due to size constraints. In this report we examine the previously published hybrid construct (H1) and a new more extensive hybrid (H2), with our documented in cellulo (live cell- based) assay utilizing a fluorescent GM2 ganglioside derivative. Unfortunately when Tay-Sachs cells were transfected with either the H1 or H2 hybrid construct and then were fed the GM2 derivative, no significant increase in its turnover was detected. In vitro assays with the isolated H1 or H2 homodimers confirmed that neither was capable of human GM2AP-dependent hydrolysis of GM2 ganglioside.

scholarly journals An α-subunit loop structure is required for GM2 activator protein binding by β-hexosaminidase A

2004 ◽  
Vol 324 (3) ◽  
pp. 1048-1052 ◽  
Author(s):  
Maryam Zarghooni ◽  
Scott Bukovac ◽  
Michael Tropak ◽  
John Callahan ◽  
Don Mahuran
Keyword(s):  
Protein Binding ◽  
Loop Structure ◽  
Α Subunit ◽  
Activator Protein ◽  
Hexosaminidase A ◽  
Gm2 Activator ◽  
Gm2 Activator Protein

Low endogenous glucocorticoid allows induction of kidney cortical cyclooxygenase-2 during postnatal rat development

2004 ◽  
Vol 286 (1) ◽  
pp. F26-F37 ◽  
Author(s):  
Kirsten Madsen ◽  
Jane Stubbe ◽  
Tianxin Yang ◽  
Ole Skøtt ◽  
Sebastian Bachmann ◽  
...  
Keyword(s):  
Kidney Development ◽  
Collecting Duct ◽  
Rat Kidney ◽  
Plasma Concentrations ◽  
Cyclooxygenase 2 ◽  
Kidney Cortex ◽  
Thick Ascending Limb ◽  
Corticosterone Concentration ◽  
Rat Kidney Cortex ◽  
Cox 2

In postnatal weeks 2–4, cyclooxygenase-2 (COX-2) is induced in the rat kidney cortex where it is critically involved in final stages of kidney development. We examined whether changes in circulating gluco- or mineralocorticosteroids or in their renal receptors regulate postnatal COX-2 induction. Plasma corticosterone concentration peaked at birth, decreased to low levels at days 3- 13, and increased to adult levels from day 22. Aldosterone peaked at birth and then stabilized at adult levels. Gluco- and mineralocorticoid receptor (GR and MR) mRNAs were expressed stably in kidney before, during, and after COX-2 induction. 11β-Hydroxysteroid dehydrogenase 2 was induced shortly after birth and was widely distributed in the whole collecting duct system in the suckling period and then returned to an adult pattern. Supplementation with corticosterone (20 mg·kg-1·day-1) or GR-specific dexamethasone (1 mg·kg-1·day-1) during low endogenous corticosterone suppressed renal COX-2 mRNA and protein and led to a restricted distribution of COX-2 immunolabeling. The ability of glucocorticoids to affect COX-2 was reflected in colocalization of GR-α and COX-2 immunoreactivity and mRNAs in thick ascending limb of Henle's loop. The MR antagonist potassium canrenoate (20 mg·kg-1·day-1) enhanced COX-2 expression from days 5 to 10, but low MR-specific concentrations of DOCA (1 mg·kg-1·day-1) had no effect on COX-2. Renomedullary interstitial cells expressed GR-α and COX-2. Dexamethasone suppressed COX-2 in these cells. Thus low plasma concentrations of corticosterone allowed for cortical and medullary COX-2 induction during postnatal kidney development. Increased circulating glucocorticoid in the postnatal period may damage late renal development through inhibition of COX-2.

scholarly journals In Cellulo Examination of a Beta-Alpha Hybrid Construct of Beta-Hexosaminidase A Subunits, Reported to Interact with the GM2 Activator Protein and Hydrolyze GM2 Ganglioside

PLoS ONE ◽  
2013 ◽  
Vol 8 (3) ◽  
pp. e57908 ◽  
Author(s):  
Incilay Sinici ◽  
Sayuri Yonekawa ◽  
Ilona Tkachyova ◽  
Steven J. Gray ◽  
R. Jude Samulski ◽  
...  
Keyword(s):  
Gm2 Ganglioside ◽  
Activator Protein ◽  
Hexosaminidase A ◽  
Hybrid Construct ◽  
Gm2 Activator ◽  
Gm2 Activator Protein

scholarly journals In Cellulo Examination of a Beta-Alpha Hybrid Construct of Beta-Hexosaminidase A Subunits, Reported to Interact with the GM2 Activator Protein and Hydrolyze GM2 Ganglioside

2021 ◽  
Author(s):  
Warren W. Wakarchuk ◽  
Incilay Sinici ◽  
Sayuri Yonekawa ◽  
Ilona Tkachyova ◽  
Steven J. Gray ◽  
...  
Keyword(s):  
In Vitro Assays ◽  
Variant Form ◽  
Gm2 Ganglioside ◽  
Activator Protein ◽  
Hexosaminidase A ◽  
Hybrid Construct ◽  
Gm2 Activator ◽  
Gm2 Activator Protein

The hydrolysis in lysosomes of GM2 ganglioside to GM3 ganglioside requires the correct synthesis, intracellular assembly and transport of three separate gene products; i.e., the alpha and beta subunits of heterodimeric beta-hexosaminidase A, E.C. # 3.2.1.52 (encoded by the HEXA and HEXB genes, respectively), and the GM2-activator protein (GM2AP, encoded by the GM2A gene). Mutations in any one of these genes can result in one of three neurodegenerative diseases collectively known as GM2 gangliosidosis (HEXA, Tay-Sachs disease, MIM # 272800; HEXB, Sandhoff disease, MIM # 268800; and GM2A, AB-variant form, MIM # 272750). Elements of both of the hexosaminidase A subunits are needed to productively interact with the GM2 ganglioside-GM2AP complex in the lysosome. Some of these elements have been predicted from the crystal structures of hexosaminidase and the activator. Recently a hybrid of the two subunits has been constructed and reported to be capable of forming homodimers that can perform this reaction in vivo, which could greatly simplify vector-mediated gene transfer approaches for Tay-Sachs or Sandhoff diseases. A cDNA encoding a hybrid hexosaminidase subunit capable of dimerizing and hydrolyzing GM2 ganglioside could be incorporated into a single vector, whereas packaging both subunits of hexosaminidase A into vectors, such as adeno-associated virus, would be impractical due to size constraints. In this report we examine the previously published hybrid construct (H1) and a new more extensive hybrid (H2), with our documented in cellulo (live cell- based) assay utilizing a fluorescent GM2 ganglioside derivative. Unfortunately when Tay-Sachs cells were transfected with either the H1 or H2 hybrid construct and then were fed the GM2 derivative, no significant increase in its turnover was detected. In vitro assays with the isolated H1 or H2 homodimers confirmed that neither was capable of human GM2AP-dependent hydrolysis of GM2 ganglioside.

Lithium-induced NDI in rats is associated with loss of α-ENaC regulation by aldosterone in CCD

2006 ◽  
Vol 290 (5) ◽  
pp. F1222-F1233 ◽  
Author(s):  
Jakob Nielsen ◽  
Tae-Hwan Kwon ◽  
Jørgen Frøkiær ◽  
Mark A. Knepper ◽  
Søren Nielsen
Keyword(s):  
Protein Expression ◽  
Collecting Duct ◽  
Rat Kidney ◽  
Lithium Treatment ◽  
Lithium Concentration ◽  
Fractional Excretion ◽  
Kidney Cortex ◽  
Cortical Collecting Duct ◽  
Rat Kidney Cortex ◽  
Fractional Excretion Of Sodium

Lithium-induced nephrogenic diabetes insipidus (Li-NDI) is associated with increased urinary sodium excretion and decreased responsiveness to aldosterone and vasopressin. Dysregulation of the epithelial sodium channel (ENaC) is thought to play an important role in renal sodium wasting. The effect of 7-day aldosterone and spironolactone treatment on regulation of ENaC in rat kidney cortex was investigated in rats with 3 wk of Li-NDI. Aldosterone treatment of rats with Li-NDI decreased fractional excretion of sodium (0.83 ± 0.02), whereas spironolactone did not change fractional excretion of sodium (1.10 ± 0.11) compared with rats treated with lithium alone (1.11 ± 0.05). Plasma lithium concentration was decreased by aldosterone (0.31 ± 0.03 mmol/l) but unchanged with spironolactone (0.84 ± 0.18 mmol/l) compared with rats treated with lithium alone (0.54 ± 0.04 mmol/l). Immunoblotting showed increased protein expression of α-ENaC, the 70-kDa form of γ-ENaC, and the Na-Cl cotransporter (NCC) in kidney cortex in aldosterone-treated rats, whereas spironolactone decreased α-ENaC and NCC compared with control rats treated with lithium alone. Immunohistochemistry confirmed increased expression of α-ENaC in the late distal convoluted tubule and connecting tubule and also revealed increased apical targeting of all three ENaC subunits (α, β, and γ) in aldosterone-treated rats compared with rats treated with lithium alone. Aldosterone did not, however, affect α-ENaC expression in the cortical collecting duct (CCD), which showed weak and dispersed labeling similar to that in rats treated with lithium alone. Spironolactone did not affect ENaC targeting compared with rats treated with lithium alone. This study shows a segment specific lack of aldosterone-mediated α-ENaC regulation in the CCD affecting both α-ENaC protein expression and trafficking, which may explain the increased sodium wasting associated with chronic lithium treatment.

scholarly journals The human GM2 activator protein. A substrate specific cofactor of beta-hexosaminidase A

1991 ◽  
Vol 266 (3) ◽  
pp. 1879-1887 ◽  
Author(s):  
E M Meier ◽  
G Schwarzmann ◽  
W Fürst ◽  
K Sandhoff
Keyword(s):  
Protein A ◽  
Activator Protein ◽  
Hexosaminidase A ◽  
Gm2 Activator ◽  
Gm2 Activator Protein

Two patients from Turkey with a novel variant in the GM2A gene and review of the literature

2021 ◽  
Vol 0 (0) ◽  
Author(s):  
Aslı İnci ◽  
Filiz Başak Cengiz Ergin ◽  
Gürsel Biberoğlu ◽  
İlyas Okur ◽  
Fatih Süheyl Ezgü ◽  
...  
Keyword(s):  
Sandhoff Disease ◽  
Pathogenic Variant ◽  
Protein Deficiency ◽  
Gm2 Gangliosidosis ◽  
Infantile Form ◽  
Rare Form ◽  
Inborn Errors ◽  
Activator Protein ◽  
Gm2 Activator ◽  
Gm2 Activator Protein

Abstract Objectives GM2 gangliosidosis is a rare form of inborn errors of metabolism including Tay-Sachs disease, Sandhoff disease, and GM2 activator deficiency. GM2 activator protein deficiency is an ultra-rare form of GM2 gangliosidosis. To date, 16 cases of GM2 activator protein deficiency have been reported in the literature, and among them, 11 cases were the infantile form of the disease. Here we report the first two patients from Turkey with the infantile form of the disease with a novel likely pathogenic variant. Case presentation A boy of eight months old presented to the metabolic department with very mild neurological deterioration, although he had achieved early developmental milestones at the appropriate time. The parents also had a daughter who had lost skills progressively before one year of age. The boy was evaluated and bilateral cherry-red spots were found with no abnormality in either metabolic screening including β-hexosaminidase or cranial magnetic resonance imaging. A novel homozygous likely pathogenic variant in GM2A was detected in a next-generation sequence panel revealing GM2 activator protein deficiency. His sister was investigated after he was diagnosed with GM2 activator deficiency and it was found that she had the same variant as her brother. Conclusions This case report emphasizes that in the event of normal β-hexosaminidase activity, GM2 activator protein deficiency could be underdiagnosed, and further molecular analysis should be performed. To the best of our knowledge, this boy is one of the youngest patient diagnosed with very mild symptoms. With this novel pathogenic variant, these patients have expanded the mutation spectrum of GM2 activator protein deficiency.

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