scholarly journals Gut microbiota regulation of P-glycoprotein in the intestinal epithelium in maintenance of homeostasis

Microbiome ◽  
2021 ◽  
Vol 9 (1) ◽  
Author(s):  
Sage E. Foley ◽  
Christine Tuohy ◽  
Merran Dunford ◽  
Michael J. Grey ◽  
Heidi De Luca ◽  
...  
Keyword(s):  
Microbial Community ◽  
Intestinal Epithelium ◽  
Critical Role ◽  
Secondary Bile Acid ◽  
Core Microbiome ◽  
P Glycoprotein ◽  
Mucosal Barrier Function ◽  
And Function

Abstract Background P-glycoprotein (P-gp) plays a critical role in protection of the intestinal epithelia by mediating efflux of drugs/xenobiotics from the intestinal mucosa into the gut lumen. Recent studies bring to light that P-gp also confers a critical link in communication between intestinal mucosal barrier function and the innate immune system. Yet, despite knowledge for over 10 years that P-gp plays a central role in gastrointestinal homeostasis, the precise molecular mechanism that controls its functional expression and regulation remains unclear. Here, we assessed how the intestinal microbiome drives P-gp expression and function. Results We have identified a “functional core” microbiome of the intestinal gut community, specifically genera within the Clostridia and Bacilli classes, that is necessary and sufficient for P-gp induction in the intestinal epithelium in mouse models. Metagenomic analysis of this core microbial community revealed that short-chain fatty acid and secondary bile acid production positively associate with P-gp expression. We have further shown these two classes of microbiota-derived metabolites synergistically upregulate P-gp expression and function in vitro and in vivo. Moreover, in patients suffering from ulcerative colitis (UC), we find diminished P-gp expression coupled to the reduction of epithelial-derived anti-inflammatory endocannabinoids and luminal content (e.g., microbes or their metabolites) with a reduced capability to induce P-gp expression. Conclusion Overall, by means of both in vitro and in vivo studies as well as human subject sample analysis, we identify a mechanistic link between cooperative functional outputs of the complex microbial community and modulation of P-gp, an epithelial component, that functions to suppress overactive inflammation to maintain intestinal homeostasis. Hence, our data support a new cross-talk paradigm in microbiome regulation of mucosal inflammation.

scholarly journals Reading and Function of a Histone Code Involved in Targeting Corepressor Complexes for Repression

2005 ◽  
Vol 25 (1) ◽  
pp. 324-335 ◽  
Author(s):  
Ho-Geun Yoon ◽  
Youngsok Choi ◽  
Philip A. Cole ◽  
Jiemin Wong
Keyword(s):  
Transcriptional Activation ◽  
Transcriptional Repression ◽  
Hormone Receptors ◽  
Critical Role ◽  
Pericentric Heterochromatin ◽  
Histone Code ◽  
Stable Association ◽  
And Function

ABSTRACT A central question in histone code theory is how various codes are recognized and utilized in vivo. Here we show that TBL1 and TBLR1, two WD-40 repeat proteins in the corepressor SMRT/N-CoR complexes, are functionally redundant and essential for transcriptional repression by unliganded thyroid hormone receptors (TR) but not essential for transcriptional activation by liganded TR. TBL1 and TBLR1 bind preferentially to hypoacetylated histones H2B and H4 in vitro and have a critical role in targeting the corepressor complexes to chromatin in vivo. We show that targeting SMRT/N-CoR complexes to the deiodinase 1 gene (D1) requires at least two interactions, one between unliganded TR and SMRT/N-CoR and the other between TBL1/TBLR1 and hypoacetylated histones. Neither interaction alone is sufficient for the stable association of the corepressor complexes with the D1 promoter. Our data support a feed-forward working model in which deacetylation exerted by initial unstable recruitment of SMRT/N-CoR complexes via their interaction with unliganded TR generates a histone code that serves to stabilize their own recruitment. Similarly, we find that targeting of the Sin3 complex to pericentric heterochromatin may also follow this model. Our studies provide an in vivo example that a histone code is not read independently but is recognized in the context of other interactions.

The microtubule modulator RanBP10 plays a critical role in regulation of platelet discoid shape and degranulation

Blood ◽  
2009 ◽  
Vol 114 (27) ◽  
pp. 5532-5540 ◽  
Author(s):  
Stefan Kunert ◽  
Imke Meyer ◽  
Silke Fleischhauer ◽  
Martin Wannack ◽  
Janine Fiedler ◽  
...  
Keyword(s):  
Fetal Liver ◽  
Critical Role ◽  
Mutant Mice ◽  
Axis Ratio ◽  
Prolonged Bleeding Time ◽  
Normal Platelet ◽  
And Function ◽  
Granule Release

Abstract Terminally mature megakaryocytes undergo dramatic cellular reorganization to produce hundreds of virtually identical platelets. A hallmark feature of this process is the generation of an elaborate system of branched protrusions called proplatelets. We recently identified RanBP10 as a tubulin-binding protein that is concentrated along polymerized microtubules in mature megakaryocytes. RanBP10 depletion in vitro caused the disturbance of polymerized filaments. Here we study the function of RanBP10 in vivo by generating deficient mice using a gene-trap approach. Mutant mice show normal platelet counts, and fetal liver-derived megakaryocytes reveal only slightly reduced proplatelet formation. However, ultrastructural analysis unveiled a significantly increased geometric axis ratio for resting platelets, and many platelets exhibited disorders in microtubule filament numbers and localization. Mutant mice showed a markedly prolonged bleeding time. Granule release, a process that depends on internal contraction of the microtubule marginal coil, also was reduced. Flow cytometry analysis revealed reduced expression of CD62P and CD63 after PAR4-peptide stimulation. These data suggest that RanBP10 plays an essential role in hemostasis and in maintaining microtubule dynamics with respect to both platelet shape and function.

scholarly journals A proline-rich sequence unique to MEK1 and MEK2 is required for raf binding and regulates MEK function.

1995 ◽  
Vol 15 (10) ◽  
pp. 5214-5225 ◽  
Author(s):  
A D Catling ◽  
H J Schaeffer ◽  
C W Reuter ◽  
G R Reddy ◽  
M J Weber
Keyword(s):  
Protein Interactions ◽  
Critical Role ◽  
Phosphorylation Site ◽  
Specific Protein ◽  
Protein Protein Interactions ◽  
Serum Stimulation ◽  
And Function

Mammalian MEK1 and MEK2 contain a proline-rich (PR) sequence that is absent both from the yeast homologs Ste7 and Byr1 and from a recently cloned activator of the JNK/stress-activated protein kinases, SEK1/MKK4. Since this PR sequence occurs in MEKs that are regulated by Raf family enzymes but is missing from MEKs and SEKs activated independently of Raf, we sought to investigate the role of this sequence in MEK1 and MEK2 regulation and function. Deletion of the PR sequence from MEK1 blocked the ability of MEK1 to associate with members of the Raf family and markedly attenuated activation of the protein in vivo following growth factor stimulation. In addition, this sequence was necessary for efficient activation of MEK1 in vitro by B-Raf but dispensable for activation by a novel MEK1 activator which we have previously detected in fractionated fibroblast extracts. Furthermore, we found that a phosphorylation site within the PR sequence of MEK1 was required for sustained MEK1 activity in response to serum stimulation of quiescent fibroblasts. Consistent with this observation, we observed that MEK2, which lacks a phosphorylation site at the corresponding position, was activated only transiently following serum stimulation. Finally, we found that deletion of the PR sequence from a constitutively activated MEK1 mutant rendered the protein nontransforming in Rat1 fibroblasts. These observations indicate a critical role for the PR sequence in directing specific protein-protein interactions important for the activation, inactivation, and downstream functioning of the MEKs.

scholarly journals Pharmacokinetic Interactions of Herbs with Cytochrome P450 and P-Glycoprotein

2015 ◽  
Vol 2015 ◽  
pp. 1-10 ◽  
Author(s):  
Hyun-Jong Cho ◽  
In-Soo Yoon
Keyword(s):  
Cytochrome P450 ◽  
Glycoprotein P ◽  
Metabolizing Enzymes ◽  
Herbal Products ◽  
P Glycoprotein ◽  
Concurrent Use ◽  
Substrate Drug ◽  
And Function

The concurrent use of drugs and herbal products is becoming increasingly prevalent over the last decade. Several herbal products have been known to modulate cytochrome P450 (CYP) enzymes and P-glycoprotein (P-gp) which are recognized as representative drug metabolizing enzymes and drug transporter, respectively. Thus, a summary of knowledge on the modulation of CYP and P-gp by commonly used herbs can provide robust fundamentals for optimizing CYP and/or P-gp substrate drug-based therapy. Herein, we review ten popular medicinal and/or dietary herbs as perpetrators of CYP- and P-gp-mediated pharmacokinetic herb-drug interactions. The main focus is placed on previous works on the ability of herbal extracts and their phytochemicals to modulate the expression and function of CYP and P-gp in severalin vitroandin vivoanimal and human systems.

scholarly journals Thymosin β4 is essential for thrombus formation by controlling the G-actin/F-actin equilibrium in platelets

Haematologica ◽  
2021 ◽  
Author(s):  
Inga Scheller ◽  
Sarah Beck ◽  
Vanessa Göb ◽  
Carina Gross ◽  
Raluca A. I. Neagoe ◽  
...  
Keyword(s):  
Thrombus Formation ◽  
Critical Role ◽  
Actin Dynamics ◽  
Thymosin Β4 ◽  
Clot Retraction ◽  
And Function ◽  
External Signals

Coordinated rearrangements of the actin cytoskeleton are pivotal for platelet biogenesis from megakaryocytes (MKs) but also orchestrate key functions of peripheral platelets in hemostasis and thrombosis, such as granule release, the formation of filopodia and lamellipodia, or clot retraction. Along with profilin (Pfn) 1, thymosin β4 (encoded by Tmsb4x) is one of the two main G-actin sequestering proteins within cells of higher eukaryotes, and its intracellular concentration is particularly high in cells that rapidly respond to external signals by increased motility, such as platelets. Here, we analyzed constitutive Tmsb4x knockout (KO) mice to investigate the functional role of the protein in platelet production and function. Thymosin β4 deficiency resulted in a macrothrombocytopenia with only mildly increased platelet volume and an unaltered platelet life span. MK numbers in the bone marrow (BM) and spleen were unaltered, however, Tmsb4x KO MKs showed defective proplatelet formation in vitro and in vivo. Thymosin β4 deficient platelets displayed markedly decreased G-actin levels and concomitantly increased F-actin levels resulting in accelerated spreading on fibrinogen and clot retraction. Moreover, Tmsb4x KO platelets showed activation defects and an impaired immunoreceptor tyrosine-based activation motif (ITAM) signaling downstream of the activating collagen receptor glycoprotein (GP) VI. These defects translated into impaired aggregate formation under flow, protection from occlusive arterial thrombus formation in vivo and increased tail bleeding times. In summary, these findings point to a critical role of thymosin β4 for actin dynamics during platelet biogenesis, platelet activation downstream of GPVI and thrombus stability.

Flt3-Ligand Plays a Critical Role in Restoring Function of NOD Facilitating Cells.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 1186-1186
Author(s):  
Yiming Huang ◽  
Isabelle J. Fugier-Vivier ◽  
Thomas Miller ◽  
Mary J. Elliott ◽  
Michael K. Tanner ◽  
...  
Keyword(s):  
Plasmacytoid Dendritic Cell ◽  
Critical Role ◽  
Nod Mice ◽  
Flt3 Ligand ◽  
Hematopoietic Stem ◽  
Functionally Impaired ◽  
And Function

Abstract CD8+/TCR− facilitating cells (FC) enhance engraftment of purified hematopoietic stem cells (HSC) in syngeneic and allogeneic recipients. FC also induce the production of regulatory T cells (Treg) in vivo and in vitro. The B220+/CD11c+/CD11b− precursor plasmacytoid dendritic cell (p-preDC) subpopulation in FC (p-preDC FC) is critical to FC function. However, p-preDC FC are significantly less efficient in function compared to FC total. In this study, we evaluated the phenotype and function of FC from diabetes-prone nonobese diabetes (NOD) mice. We found that NOD FC contain subpopulations similar to those previously described in B6 FC, including p-preDC, CD19+, NK1.1+DX5+ and myeloid cells. P-preDC represent the major FC subpopulation in NOD mice. The CD19+, DX5+ and B220−/CD11c+/CD11b+ subpopulations were significantly decreased in NOD FC compared to those from B6 or NOR mice (Figure 1a; * = P< 0.05; ** = P< 0.007). To test the function of NOD FC, 500 HSC (c-Kit+/Sca-1+/Lin−) were sorted and transplanted with or without 30,000 FC into conditioned 950 cGy recipients. MHC-matched diabetes-resistant NOR mice were served as a control strain for NOD mice. 5 (31%) of 16 recipients of NOR HSC were engrafted and survived up to 130 days. 7 (70%) of 10 recipients of NOR HSC plus FC have long-term engraftment and survival over 130 days, indicating that the NOR FC significantly enhanced engraftment of NOR HSC compared to HSC alone. In striking contrast, NOD FC were functionally impaired and did not enhance HSC engraftment in NOD recipients as evidenced by similar engraftment of HSC with FC (31%, n = 13) compared to the HSC alone (24%, n = 17; P = 0.579). Notably, when NOD mice were treated with Flt3 ligand (FL; 10 μg/ subcutaneous/daily, 10 days), FC were expanded in peripheral blood (PB). The DX5+ and B220−/CD11c+/CD11b+ subpopulations were significantly increased (Figure 1b). The FL-PB FC significantly facilitate engraftment of allogeneic HSC in vivo in the NOD → B10 model (Figure 1c; P = 0.02). These data demonstrate that NOD FC exhibit significantly impaired function and FL plays an important role in regulation and development of FC function. We propose that the defective function of NOD FC is mechanistically due either to the abnormal activation status of the p-preDC FC population and/or the absence of collaborative subpopulations in FC such as NK FC. This hypothesis offers an attractive explanation for the mechanism of FC to enhance HSC engraftment and to induce tolerance. Studies are underway to evaluate the mechanism by which FC contribute to diabetes-pathogenesis and/or prevention. Figure Figure

scholarly journals Monocrotaline Suppresses RANKL-Induced Osteoclastogenesis In Vitro and Prevents LPS-Induced Bone Loss In Vivo

2018 ◽  
Vol 48 (2) ◽  
pp. 644-656 ◽  
Author(s):  
Cheng-Ming Wei ◽  
Yi-Ji Su ◽  
Xiong Qin ◽  
Jia-Xin Ding ◽  
Qian Liu ◽  
...  
Keyword(s):  
Critical Role ◽  
Osteoclast Differentiation ◽  
Osteoclast Formation ◽  
Bone Diseases ◽  
Marker Genes ◽  
Tartrate Resistant Acid Phosphatase ◽  
Dependent Manner ◽  
And Function

Background/Aims: Extensive osteoclast formation plays a critical role in bone diseases, including rheumatoid arthritis, periodontitis and the aseptic loosening of orthopedic implants. Thus, identification of agents that can suppress osteoclast formation and bone resorption is important for the treatment of these diseases. Monocrotaline (Mon), the major bioactive component of crotalaria sessiliflora has been investigated for its anti-cancer activities. However, the effect of Mon on osteoclast formation and osteolysis is not known. Methods: The bone marrow macrophages (BMMs) were cultured with M-CSF and RANKL followed by Mon treatment. Then the effects of Mon on osteoclast differentiation were evaluated by counting TRAP (+) multinucleated cells. Moreover, effects of Mon on hydroxyapatite resorption activity of mature osteoclast were studied through resorption areas measurement. The involved potential signaling pathways were analyzed by performed Western blotting and quantitative real-time PCR examination. Further, we established a mouse calvarial osteolysis model to measure the osteolysis suppressing effect of Mon in vivo. Results: In this study, we show that Mon can inhibit RANKL-induced osteoclast formation and function in a dose-dependent manner. Mon inhibits the expression of osteoclast marker genes such as tartrate-resistant acid phosphatase (TRAP) and cathepsin K. Furthermore, Mon inhibits RANKL-induced the activation of p38 and JNK. Consistent with in vitro results, Mon exhibits protective effects in an in vivo mouse model of LPS-induced calvarial osteolysis. Conclusion: Taken together our data demonstrate that Mon may be a potential prophylactic anti-osteoclastic agent for the treatment of osteolytic diseases caused by excessive osteoclast formation and function.

scholarly journals The role of Mfn2 in the structure and function of ER-mitochondrial tethering in vivo

2021 ◽  
Author(s):  
Song Han ◽  
Fanpeng Zhao ◽  
Jeffrey Hsia ◽  
Xiaopin Ma ◽  
Yi Liu ◽  
...  
Keyword(s):  
Pyramidal Neurons ◽  
Critical Role ◽  
Biochemical Changes ◽  
Conditional Knockout ◽  
Functional Studies ◽  
And Function ◽  
Close Contacts

The mitochondria-ER contacts (MERCs) plays an essential role in multiple cell physiological process. While Mfn2 was the first protein implicated in the formation of MERCs, it is debated whether it acts as a tether or antagonizer, largely based on in vitro studies. To understand the role of Mfn2 in MERCs in vivo, we characterized ultrastructural and biochemical changes of MERCs in pyramidal neurons of hippocampus in Mfn2 conditional knockout (KO) mice and in Mfn2 overexpression (OE) mice and found Mfn2 ablation caused reduced close contacts while Mfn2 OE caused increased close contacts between ER and mitochondria in vivo. Functional studies on SH-SY5Y cells with Mfn2 KO or overexpression demonstrating similar biochemical changes found that mitochondrial calcium uptake along with IP3R3-Grp75 interaction was decreased in Mfn2 KO cells but increased in the Mfn2 OE cells. Lastly, we found Mfn2 KO decreased and Mfn2 OE increased the interaction between the ER-mitochondria tethering pair of VAPB-PTPIP51. In conclusion, our study supports the notion that Mfn2 plays a critical role in ER-mitochondrial tethering and the formation of close contacts in neuronal cells in vivo.

scholarly journals Septin-7 is indispensable for proper skeletal muscle architecture and function

2021 ◽  
Author(s):  
Monika Gonczi ◽  
Zsolt Raduly ◽  
Laszlo Szabo ◽  
Janos Fodor ◽  
Andrea Telek ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Critical Role ◽  
C2c12 Cell ◽  
C2c12 Cells ◽  
Knock Out ◽  
Fourth Component ◽  
And Function ◽  
Mitochondrial Networks

Today septins are considered as the fourth component of the cytoskeleton with the Septin-7 isoform playing a critical role in the formation of higher order structures. While its importance has already been confirmed in several intracellular processes of different organs, very little is known about its role in skeletal muscle. Here, using Septin-7 conditional knock-down mouse model, the C2C12 cell line, and enzymatically isolated adult muscle fibers the organization and localization of septin filaments is revealed, and an ontogenesis-dependent expression of Septin-7 is demonstrated. KD mice displayed a characteristic hunchback phenotype with skeletal deformities, reduction in vivo and in vitro force generation, and disorganized mitochondrial networks. Furthermore, knock-out of Septin-7 in C2C12 cells resulted in complete loss of cell division while KD cells provided evidence that Septin-7 is essential in proper myotube differentiation. These and the transient increase in Septin-7 expression following muscle injury demonstrate its vital contribution to muscle regeneration and development.

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