Megakaryocytic Maturation in Response to Shear Flow Is Mediated by the Activator Protein 1 (AP-1) Transcription Factor via Mitogen-activated Protein Kinase (MAPK) Mechanotransduction

Luff, Stephanie A.; Papoutsakis, Eleftherios T.

doi:10.1074/jbc.m115.707174

Cited by 25 publications

(27 citation statements)

References 77 publications

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“…In addition, NF-κB activation in the airways of allergen-challenged mice is attenuated by Toll-like receptor (TLR) 2 or TLR4 gene deletion, suggesting that TLR2 or TLR4 contribute to NF-κB signaling (16,17). Similarly, the AP-1 and STAT3 promoter-binding activity are regulated by MAPK and JAK/STAT signaling (18,19). Therefore, it was hypothesized that LPS may induce NF-κB, MAPK and JAK/STAT pathway activation, leading to NF-κB, AP-1 and STAT3 nuclear translocation and cytokine expression in the H292 and THP-1 cells investigated in the present study.…”

Section: Effect Of Lps Treatment On Nf-κb Ap-1 Ppar and Stat3mentioning

confidence: 99%

LPS‑induced proinflammatory cytokine expression in human airway epithelial cells and macrophages via NF‑κB, STAT3 or AP‑1 activation

et al. 2018

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Lipopolysaccharide (LPS), the major outer surface membrane component of Gram-negative bacteria, is one of the main etiological factors in the pathogenesis of several lung diseases, such as chronic obstructive pulmonary disease. The respiratory epithelium and the macrophages comprise the dynamic interface between the outside environment and the host response to bacterial infection via cytokine secretion. In the present study, the mechanisms of LPS induced‑inflammatory response in human lung cells and macrophages were investigated. The effects of LPS exposure on cytokine production, inflammation‑related transcription factors and intracellular signaling pathway activation were assessed in human lung mucoepidermoid carcinoma H292 cells and human macrophage THP‑1 cells. The results demonstrated that LPS markedly increased the expression of interleukin (IL)‑6, IL‑8, tumor necrosis factor (TNF)‑α, matrix metallopeptidase (MMP)‑9 and tissue inhibitor of metalloproteinases‑1 in H292 cells, while it increased the production of IL‑6, IL‑8 and TNF‑α in differentiated THP‑1 cells. In addition, LPS exposure activated nuclear factor (NF)‑κB and activator protein (AP)‑1 signaling in H292 cells, while it activated NF‑κB and signal transducer and activator of transcription (STAT) 3 signaling in THP‑1 cells. Furthermore, treatment with NF‑κB, AP‑1 or STAT3 inhibitors significantly decreased the LPS‑mediated expression of IL‑8 and TNF‑α in these cells, suggesting that these pathways might serve crucial roles in LPS‑induced cytokine expression. In conclusion, LPS stimulation of H292 and THP‑1 cells induced cytokine expression and NF‑κB, mitogen‑activated protein kinase and Janus kinase/STAT3 pathway activation with subsequent nuclear translocation of NF‑κB, AP‑1 and STAT3, which demonstrated potential of the use of NF‑κB, AP‑1 and STAT3 in therapies for conditions and diseases associated with chronic inflammation.

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Section: Effect Of Lps Treatment On Nf-κb Ap-1 Ppar and Stat3mentioning

confidence: 99%

LPS‑induced proinflammatory cytokine expression in human airway epithelial cells and macrophages via NF‑κB, STAT3 or AP‑1 activation

et al. 2018

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“…Fibrinogen mediated adhesion and signaling is an important component of megakaryocyte differentiation, development, and proplatelet formation . Fibrinogen signaling also activates translation control pathways, including the PI3K and MAPK pathways . As noted above, in other cells PDK1 regulates two pathways that converge on translation: first, Akt, which activates mTORC1 mediated translation and second, ERK1/2, which activates MAPK mediated translation .…”

Section: Resultsmentioning

confidence: 97%

Phospho‐inositide‐dependent kinase 1 regulates signal dependent translation in megakaryocytes and platelets

Manne

Bhatlekar

Middleton

et al. 2020

Journal of Thrombosis and Haemostasis

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Background: Regulated protein synthesis is essential for megakaryocyte (MK) and platelet functions, including platelet production and activation. PDK1 (phosphoinositide-dependent kinase 1) regulates platelet functional responses and has been associated with circulating platelet counts. Whether PDK1 also directly regulates protein synthetic responses in MKs and platelets, and platelet production by MKs, remains unknown. Objective: To determine if PDK1 regulates protein synthesis in MKs and platelets. Methods: Pharmacologic PDK1 inhibitors (BX-795) and mice where PDK1 was selectively ablated in MKs and platelets (PDK1 −/− ) were used. PDK1 signaling in MKs and platelets (human and murine) were assessed by immunoblots. Activation-dependent translation initiation and protein synthesis in MKs and platelets was assessed by probing for dissociation of eIF4E from 4EBP1, and using m7-GTP pulldowns and S 35 methionine incorporation assays. Proplatelet formation by MKs, synthesis of Bcl-3 and MARCKs protein, and clot retraction were employed for functional assays. Results: Inhibiting or ablating PDK1 in MKs and platelets abolished the phosphorylation of 4EBP1 and eIF4E by preventing activation of the PI3K and MAPK pathways. Inhibiting PDK1 also prevented dissociation of eIF4E from 4EBP1, decreased binding of eIF4E to m7GTP (required for translation initiation), and significantly reduced de novo protein synthesis. Inhibiting PDK1 reduced proplatelet formation by human MKs and blocked MARCKs protein synthesis. In both human and murine platelets, PDK1 controlled Bcl-3 synthesis. Inhibition of PDK1 led to complete failure of clot retraction in vitro. Conclusions: PDK1 is a previously unidentified translational regulator in MKs and platelets, controlling protein synthetic responses, proplatelet formation, and clot retraction. K E Y W O R D S platelet activation, platelets, protein translation, signal transduction, thrombosis S U PP O RTI N G I N FO R M ATI O N Additional supporting information may be found online in the Supporting Information section. How to cite this article: Manne BK, Bhatlekar S, Middleton EA, Weyrich AS, Borst O, Rondina MT. Phospho-inositidedependent kinase 1 regulates signal dependent translation in megakaryocytes and platelets. J Thromb Haemost.

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“…56 We have also demonstrated that JNK signaling is involved in shear-induced Mk maturation and platelet production. 57 mTOR is a major regulator of Mk development and maturation. 58 Here, we showed that CD41 expression was significantly lower at days 7 and 12 in cells treated with an mTOR inhibitor prior to co-culture with MkMPs ( Figure 5A & 5B).…”

Section: -5p Inducesmentioning

confidence: 99%

Combinatorial effect of miR-486-5p & miR-22-3p mimics thrombopoietin’s impact on hematopoietic stem & progenitor cells

Kao

Jiang

Papoutsakis

2020

Preprint

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Word Count (Text): 5429 Word Count (Abstract): 246 Figure Count: 7Table Count: 2 Reference Count: 61 Key Points• miR-486-5p and miR-22-3p drive megakaryocytic differentiation in the absence of thrombopoietin.• Megakaryocytic microparticles trigger megakaryocytic differentiation of CD34 + cells through JNK and PI3K/Akt/mTOR signaling. AbstractMegakaryocytes shed and release submicron size microparticles (MkMPs), the most abundant microparticle in circulation. We have previously reported that MkMPs target peripheral-blood CD34 + hematopoietic stem/progenitor cells (HSPCs) to induce megakaryocytic differentiation and proliferation, and that small RNAs delivered to HSPCs via MkMPs play an important role in the development of this phenotype. Here, using single-molecule real-time (SMRT) RNA sequencing (RNAseq), we identify the top seven most abundant microRNAs (miRs) in MkMPs as potential candidates in mediating the effect of MkMPs on HSPCs. Using miR mimics, we demonstrate that among the seven most abundant miRs, two, miR-486-5p and miR-22-3p, are able to drive the Mk differentiation of HSPCs in the absence of thrombopoietin (TPO). The effect of these two miRs is comparable to the TPO-or MkMP-induced megakaryocytic differentiation of HSPCs, thus suggesting that these two miRs are responsible for this MkMPinduced phenotype. To probe the signaling through which MkMPs might enable this phenotype, we used kinase inhibitors of potential signaling pathways engaged in megakaryocytic differentiation. Our data suggest that MkMP-induced Mk differentiation of HSPCs is enabled through JNK and PI3K/Akt/mTOR signaling. Our data show that MkMPs activate Akt and mTOR phosphorylation. Furthermore, MkMPs downregulate PTEN expression, a direct target of miR-486-5p and a negative regulator of PI3K/Akt signaling, via JNK signaling. Taken together, our data provide a mechanistic understanding of the biological effect of MkMPs in inducing megakaryocytic differentiation of HSPCs, which, as was previously suggested, is a phenotype of potential physiological significance in stress megakaryopoiesis.

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Megakaryocytic Maturation in Response to Shear Flow Is Mediated by the Activator Protein 1 (AP-1) Transcription Factor via Mitogen-activated Protein Kinase (MAPK) Mechanotransduction

Cited by 25 publications

References 77 publications

LPS‑induced proinflammatory cytokine expression in human airway epithelial cells and macrophages via NF‑κB, STAT3 or AP‑1 activation

LPS‑induced proinflammatory cytokine expression in human airway epithelial cells and macrophages via NF‑κB, STAT3 or AP‑1 activation

Phospho‐inositide‐dependent kinase 1 regulates signal dependent translation in megakaryocytes and platelets

Combinatorial effect of miR-486-5p & miR-22-3p mimics thrombopoietin’s impact on hematopoietic stem & progenitor cells

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