Nongenomic STAT5-dependent effects on Golgi apparatus and endoplasmic reticulum structure and function

Lee, Jason E.; Yang, Yang‐Ming; Liang, Feng-Xia; Gough, Daniel J.; Levy, David E.; Sehgal, Pravin B.

doi:10.1152/ajpcell.00379.2011

Cited by 49 publications

(115 citation statements)

References 41 publications

(92 reference statements)

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“…For a variety of reasons (28,63,64), we initiated an investigation into the involvement of STAT5a/b species in the pathogenesis of PH produced in mice exposed to chronic hypoxia. Mice with heterozygous or homozygous conditional deletions of the STAT5a/b locus in vascular SMCs were generated in crosses between STAT5a/b fl/fl and transgelin (SM22α)-Cre +/+ parents (28) ( Figure 5).…”

Section: A Novel Starting Point For Our Stat5 Studies On Sex Bias In Phmentioning

confidence: 99%

Hypothesis: Neuroendocrine Mechanisms (Hypothalamus-Growth Hormone-STAT5 Axis) Contribute to Sex Bias in Pulmonary Hypertension

Sehgal

Yang

Miller

2015

Mol Med

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Pulmonary hypertension (PH) is a disease with high morbidity and mortality. The prevalence of idiopathic pulmonary arterial hypertension (IPAH) and hereditary pulmonary arterial hypertension (HPAH) is approximately two-to four-fold higher in women than in men. Paradoxically, there is an opposite male bias in typical rodent models of PH (chronic hypoxia or monocrotaline); in these models, administration of estrogenic compounds (for example, estradiol-17β [E2]) is protective. Further complexities are observed in humans ingesting anorexigens (female bias) and in rodent models, such as after hypoxia plus SU5416/Sugen (little sex bias) or involving serotonin transporter overexpression or dexfenfluramine administration (female bias). These complexities in sex bias in PH remain incompletely understood. We recently discovered that conditional deletion of signal transducer and activator of transcription 5a/b (STAT5a/b) in vascular smooth muscle cells abrogated the male bias in PH in hypoxic mice and that late-stage obliterative lesions in patients of both sexes with IPAH and HPAH showed reduced STAT5a/b, reduced Tyr-P-STAT5 and reduced B-cell lymphoma 6 protein (BCL6). In trying to understand the significance of these observations, we realized that there existed a wellcharacterized E2-sensitive central neuroendocrine mechanism of sex bias, studied over the last 40 years, that, at its peripheral end, culminated in species-specific male ("pulsatile") versus female ("more continuous") temporal patterns of circulating growth hormone (GH) levels leading to male versus female patterned activation of STAT5a/b in peripheral tissues and thus sex-biased expression of hundreds of genes. In this report, we consider the contribution of this neuroendocrine mechanism (hypothalamus-GH-STAT5) in the generation of sex bias in different PH situations.

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Section: A Novel Starting Point For Our Stat5 Studies On Sex Bias In Phmentioning

confidence: 99%

Hypothesis: Neuroendocrine Mechanisms (Hypothalamus-Growth Hormone-STAT5 Axis) Contribute to Sex Bias in Pulmonary Hypertension

Sehgal

Yang

Miller

2015

Mol Med

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“…19,20 The cells were imaged using a Zeiss AxioImager M2 motorized microscopy system with Zeiss W N-Achroplan 40×/NA0.75 or Zeiss EC Plan-Neofluor 100×/NA1.3 oil objectives equipped with a high-resolution RGB HRc AxioCam camera and AxioVision software (ver. 4.8.1) in 1,388 × 1,040-pixel high-speed color-capture mode.…”

Section: Temperature-sensitive Vsv-g-gfp Trafficking Assaymentioning

confidence: 99%

“…row) and subsequent appearance of the VSV-G at the cell surface (Fig. 1A, bottom row), as assayed by surface binding of the 8G5 mAb to live cells in the cold (4˚C; to stop endocytosis of the mAb 19,34 ). Analyses of the extent of trafficking in terms of the ratio of cell-surface red immunofluorescence to total VSV-G-GFP green fluorescence provided a quantitative measure of the kinetics of VSV-G trafficking from the ER to the Golgi apparatus and from there to the plasma membrane ( Fig.…”

Section: Mctp Blocks Tso45vsv-g-gfp Trafficking To the Surface Of Endmentioning

confidence: 99%

“…The trafficking data in Figure 4A and 4B confirm the previous observation that the combination of STAT5a/b downregulation reduced VSV-G trafficking. 19 We then investigated the effects of combining eNOS or BMPR2 downregulation with downregulation of STAT5a or STAT5b. The data in Figure 5A and 5B show that knockdown of endogenous eNOS significantly enhanced the trafficking defect produced by STAT5a or STAT5b downregulation alone.…”

Section: Mctp Blocks Tso45vsv-g-gfp Trafficking To the Surface Of Endmentioning

confidence: 99%

“…We discovered this "nongenomic" membrane trafficking function in the cytoplasm 19,20 for the "transcription factors" STAT5a and STAT5b. [21][22][23][24] STAT5a/b proteins associated with structural components of the ER/Golgi and their downregulation led to a characteristic cystic ER/Golgi fragmentation phenotype in primary human pulmonary arterial endothelial cells (HPAECs) and human pulmonary arterial smooth muscle cells (HPASMCs).…”

Section: Introductionmentioning

confidence: 99%

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Subcellular Mechanisms in Pulmonary Arterial Hypertension: Combinatorial Modalities that Inhibit Anterograde Trafficking and Cause Bone Morphogenetic Protein Receptor Type 2 Mislocalization

2013

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Abstract:The natural history of familial pulmonary arterial hypertension (PAH) typically involves mutations in and/or haploinsuffciency of BMPR2 (gene for bone morphogenetic protein receptor type 2) but with low penetrance (10%-15%), delayed onset (in the third or fourth decade), and a gender bias (two-to fourfold more prevalent in postpubertal women). Thus, investigators have sought an understanding of "second-hit" modalities that might affect BMPR2 anterograde trafficking and/or function. Indeed, vascular lung lesions in PAH have been reported to contain enlarged "vacuolated" endothelial and smooth muscle cells with dilated endoplasmic reticulum (ER) cisternae, increased ER structural protein reticulon 4 (also called Nogo-B), and enlarged and fragmented Golgi apparatus. We recently replicated this cellular phenotype in primary human pulmonary arterial endothelial cells and human pulmonary arterial smooth muscle cells in culture by acute knockdown of the estradiol 17β (E2)-responsive proteins signal transducer and activator of transcription 5a (STAT5a) and STAT5b using small interfering RNAs (siRNAs). We have now investigated whether functional haploinsufficiences of these molecules, alone or in combination with other modalities, might interfere with anterograde membrane trafficking using (a) the quantitative tsO45VSV-G-GFP trafficking assay and (b) assays for cell-surface localization of Flagtagged BMPR2 molecules. The G glycoprotein of the vesicular stomatitis virus (VSV-G) trafficking assay was validated in EA.hy926 endothelial cells by showing that cells exposed to monocrotaline pyrrole displayed reduced anterograde trafficking. Thereafter, the combinatorial knockdowns of STAT5a, STAT5b, BMPR2, and/or endothelial nitric oxide synthase as well as exposure to E2 or 2-methoxyestradiol were observed to significantly inhibit VSV-G trafficking. These combinations also led to intracellular trapping of wild-type Flag-tagged BMPR2. Overexpression of the PAH disease-derived F14 and KDF mutants of BMPR2, which were trapped in the ER/Golgi, also inhibited VSV-G trafficking in trans. zation of the KDF but not the F14 mutant. These data identify several combinatorial modalities that inhibit VSV-G anterograde trafficking and cause mislocalization of BMPR2. These modalities merit consideration in defining aspects of the late-developing and gender-biased natural history of human PAH.Keywords: vascular cell remodeling, membrane protein trafficking, tsO45VSV-G-GFP trafficking assay, monocrotaline pyrrole (MCTP), signal transducer and activator of transcription 5a/b (STAT5a/b), endothelial nitric oxide synthase (eNOS), estradiol 17β, probenecid.

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STATs in Health and Disease

Liongue

Lewis

Ward

2016

Cancer Drug Discovery and Development

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Nongenomic STAT5-dependent effects on Golgi apparatus and endoplasmic reticulum structure and function

Cited by 49 publications

References 41 publications

Hypothesis: Neuroendocrine Mechanisms (Hypothalamus-Growth Hormone-STAT5 Axis) Contribute to Sex Bias in Pulmonary Hypertension

Hypothesis: Neuroendocrine Mechanisms (Hypothalamus-Growth Hormone-STAT5 Axis) Contribute to Sex Bias in Pulmonary Hypertension

Subcellular Mechanisms in Pulmonary Arterial Hypertension: Combinatorial Modalities that Inhibit Anterograde Trafficking and Cause Bone Morphogenetic Protein Receptor Type 2 Mislocalization

STATs in Health and Disease

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