scholarly journals Reversing the miRNA -5p/-3p stoichiometry reveals physiological roles and targets of miR-140 miRNAs

2021 ◽  
Author(s):  
Cameron J Young ◽  
Melissa Caffrey ◽  
Christopher Janton ◽  
Tatsuya Kobayashi
Keyword(s):  
Untranslated Region ◽  
Bone Growth ◽  
Bone Development ◽  
Sequencing Analysis ◽  
Endochondral Bone ◽  
Chondrocyte Maturation ◽  
Protein Loading ◽  
Argonaute Proteins

The chondrocyte specific miR-140 miRNAs are necessary for normal endochondral bone growth in mice. miR-140 deficiency causes dwarfism and craniofacial deformity. However, the physiologically important targets of miR-140 miRNAs are still unclear. The miR-140 gene (Mir140) encodes three chondrocyte-specific microRNAs, miR-140-5p, derived from the 5′ strand of primary miR-140, and miR140-3p.1 and -3p.2, derived from the 3′ strand of primary miR-140. miR-140-3p miRNAs are ten times more abundant than miR-140-5p likely due to the non-preferential loading of miR-140-5p to Argonaute proteins. To differentiate the role of miR-140-5p and -3p miRNAs in endochondral bone development, two distinct mouse models, miR140-C>T, in which the first nucleotide of miR-140-5p was altered from cytosine to uridine, and miR140-CG, where the first two nucleotides of miR-140-3p were changed to cytosine and guanine, were created. These changes are expected to alter Argonaute protein loading preference of -5p and -3p to increase -5p loading and decrease -3p loading without changing the function of miR140-5p. These models presented a mild delay in epiphyseal development with delayed chondrocyte maturation. Using RNA-sequencing analysis of the two models, direct targets of miR140-5p, including Wnt11, were identified. Disruption of the predicted miR140-5p binding site in the 3′ untranslated region of Wnt11 was shown to increase Wnt11 mRNA expression and caused a modest acceleration of epiphyseal development. These results show that the relative abundance of miRNA-5p and -3p can be altered by changing the first nucleotide of miRNAs in vivo, and this method can be useful to identify physiologically important miRNA targets.

scholarly journals Rac signaling in osteoblastic cells is required for normal bone development but is dispensable for hematopoietic development

Blood ◽  
2012 ◽  
Vol 119 (3) ◽  
pp. 736-744 ◽  
Author(s):  
Steven W. Lane ◽  
Serena De Vita ◽  
Kylie A. Alexander ◽  
Ruchan Karaman ◽  
Michael D. Milsom ◽  
...  
Keyword(s):  
Bone Growth ◽  
Bone Development ◽  
Long Bone ◽  
Perivascular Niche ◽  
Hematopoietic Stem ◽  
Hsc Niche ◽  
Rac1 Gtpase

Abstract Hematopoietic stem cells (HSCs) interact with osteoblastic, stromal, and vascular components of the BM hematopoietic microenvironment (HM) that are required for the maintenance of long-term self-renewal in vivo. Osteoblasts have been reported to be a critical cell type making up the HSC niche in vivo. Rac1 GTPase has been implicated in adhesion, spreading, and differentiation of osteoblast cell lines and is critical for HSC engraftment and retention. Recent data suggest a differential role of GTPases in endosteal/osteoblastic versus perivascular niche function. However, whether Rac signaling pathways are also necessary in the cell-extrinsic control of HSC function within the HM has not been examined. In the present study, genetic and inducible models of Rac deletion were used to demonstrate that Rac depletion causes impaired proliferation and induction of apoptosis in the OP9 cell line and in primary BM stromal cells. Deletion of Rac proteins caused reduced trabecular and cortical long bone growth in vivo. Surprisingly, HSC function and maintenance of hematopoiesis in vivo was preserved despite these substantial cell-extrinsic changes. These data have implications for therapeutic strategies to target Rac signaling in HSC mobilization and in the treatment of leukemia and provide clarification to our evolving concepts of HSC-HM interactions.

scholarly journals PD-1 expression on NK cells can be related to cytokine stimulation and tissue residency

2021 ◽  
Author(s):  
Arnika K Wagner ◽  
Nadir Kadri ◽  
Chris Tibbitt ◽  
Koen van de Ven ◽  
Sunitha Bagawath-Singh ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
T Cells ◽  
Nk Cells ◽  
Mhc Class I ◽  
Sequencing Analysis ◽  
Single Cell Rna Sequencing ◽  
Cytokine Stimulation ◽  
Deficient Mice

ABSTRACTAlthough PD-1 was shown to be a hallmark of T cells exhaustion, controversial studies have been reported on the role of PD-1 on NK cells. Here, we found by flow cytometry and single cell RNA sequencing analysis that PD-1 can be expressed on MHC class I-deficient tumor-infiltrating NK cells in vivo. We also demonstrate distinct alterations in the phenotype of PD-1-deficient NK cells which in part could be attributed to a decrease in tumor-infiltrating NK cells in PD-1-deficient mice. NK cells from PD-1-deficient mice exhibited a more mature phenotype which might reduce their capacity to migrate and kill in vivo. Finally, our results demonstrate that PD-L1 molecules in membranes of PD-1-deficient NK cells migrate faster than in NK cells from wildtype mice, suggesting that PD-1 and PD-L1 form cis interactions with each other on NK cells.

Control of chondrocyte gene expression by actin dynamics: a novel role of cholesterol/Ror-α signalling in endochondral bone growth

2010 ◽  
Vol 13 (9b) ◽  
pp. 3497-3516 ◽  
Author(s):  
Anita Woods ◽  
Claudine G. James ◽  
Guoyan Wang ◽  
Holly Dupuis ◽  
Frank Beier
Keyword(s):  
Gene Expression ◽  
Bone Growth ◽  
Actin Dynamics ◽  
Endochondral Bone

Role of the sodium-dependent phosphate transporter PIT1 in bone growth in vivo

Bone ◽  
2011 ◽  
Vol 48 ◽  
pp. S107
Author(s):  
A. Bourgine ◽  
P. Pilet ◽  
S. Diouani ◽  
S. Sourice ◽  
J. Lesoeur ◽  
...  
Keyword(s):  
Bone Growth ◽  
Phosphate Transporter ◽  
Sodium Dependent

scholarly journals The Mammalian Target of Rapamycin Complex 1 Regulates Leptin Biosynthesis in Adipocytes at the Level of Translation: The Role of the 5′-Untranslated Region in the Expression of Leptin Messenger Ribonucleic Acid

2008 ◽  
Vol 22 (10) ◽  
pp. 2260-2267 ◽  
Author(s):  
Partha Chakrabarti ◽  
Takatoshi Anno ◽  
Brendan D. Manning ◽  
Zhijun Luo ◽  
Konstantin V. Kandror
Keyword(s):  
Untranslated Region ◽  
Molecular Mechanisms ◽  
Mammalian Target Of Rapamycin ◽  
Dominant Negative ◽  
Target Of Rapamycin ◽  
Messenger Ribonucleic Acid ◽  
Leptin Expression ◽  
Adipose Cells

Abstract Leptin production by adipose cells in vivo is increased after feeding and decreased by food deprivation. However, molecular mechanisms that control leptin expression in response to food intake remain unknown. Here, we test the hypothesis that leptin expression in adipose cells is regulated by nutrient- and insulin-sensitive mammalian target of rapamycin complex 1 (mTORC1)-mediated pathway. The activity of mTORC1 in 3T3-L1 adipocytes was up-regulated by stable expression of either constitutively active Rheb or dominant-negative AMP-activated protein kinase. In both cases, expression of endogenous leptin was significantly elevated at the level of translation. To investigate the role of leptin 5′-untranslated region (UTR) in the regulation of protein expression, we created bicistronic reporter constructs with and without the 5′-UTR. We found that the presence of leptin 5′-UTR renders mRNA resistant to regulation by mTORC1. It appears, therefore, that mTORC1 controls translation of leptin mRNA via a novel mechanism that does not require the presence of either the 5′-terminal oligopyrimidine tract or the 5′-UTR.

scholarly journals Mice Lacking the Calcineurin Inhibitor Rcan2 Have an Isolated Defect of Osteoblast Function

Endocrinology ◽  
2012 ◽  
Vol 153 (7) ◽  
pp. 3537-3548 ◽  
Author(s):  
J. H. Duncan Bassett ◽  
John G. Logan ◽  
Alan Boyde ◽  
Moira S. Cheung ◽  
Holly Evans ◽  
...  
Keyword(s):  
T Cells ◽  
Bone Development ◽  
Skeletal Development ◽  
Dominant Negative ◽  
Nuclear Factor ◽  
Activated T Cells ◽  
Osteoblast Function ◽  
And Function

Calcineurin-nuclear factor of activated T cells signaling controls the differentiation and function of osteoclasts and osteoblasts, and regulator of calcineurin-2 (Rcan2) is a physiological inhibitor of this pathway. Rcan2 expression is regulated by T3, which also has a central role in skeletal development and bone turnover. To investigate the role of Rcan2 in bone development and maintenance, we characterized Rcan2−/− mice and determined its skeletal expression in T3 receptor (TR) knockout and thyroid-manipulated mice. Rcan2−/− mice had normal linear growth but displayed delayed intramembranous ossification, impaired cortical bone formation, and reduced bone mineral accrual during development as well as increased mineralization of adult bone. These abnormalities resulted from an isolated defect in osteoblast function and are similar to skeletal phenotypes of mice lacking the type 2 deiodinase thyroid hormone activating enzyme or with dominant-negative mutations of TRα, the predominant TR isoform in bone. Rcan2 mRNA was expressed in primary osteoclasts and osteoblasts, and its expression in bone was differentially regulated in TRα and TRβ knockout and thyroid-manipulated mice. However, in primary osteoblast cultures, T3 treatment did not affect Rcan2 mRNA expression or nuclear factor of activated T cells c1 expression and phosphorylation. Overall, these studies establish that Rcan2 regulates osteoblast function and its expression in bone is regulated by thyroid status in vivo.

scholarly journals Cartilage Ablation of Sirt1 Causes Inhibition of Growth Plate Chondrogenesis by Hyperactivation of mTORC1 Signaling

Endocrinology ◽  
2019 ◽  
Vol 160 (12) ◽  
pp. 3001-3017 ◽  
Author(s):  
Xinxin Jin ◽  
Xiaomin Kang ◽  
Liting Zhao ◽  
Mao Xu ◽  
Tianping Xie ◽  
...  
Keyword(s):  
Growth Plate ◽  
Growth Retardation ◽  
Bone Growth ◽  
Bone Development ◽  
Conditional Knockout ◽  
Negative Regulator ◽  
Sirtuin 1 ◽  
Growth Plate Chondrocytes ◽  
Mtorc1 Signaling

Abstract A growing body of evidence implies a pivotal role of sirtuin-1 (Sirt1) in chondrocyte function and homeostasis; however, its underlying mechanisms mediating chondrogenesis, which is an essential process for physiological skeletal growth, are still poorly understood. In the current study, we generated TamCartSirt1−/− [Sirt1 conditional knockout (cKO)] mice to explore the role of Sirt1 during postnatal endochondral ossification. Compared with control mice, cKO mice exhibited growth retardation associated with inhibited chondrocyte proliferation and hypertrophy, as well as activated apoptosis. These effects were regulated by hyperactivation of mammalian target of rapamycin complex 1 (mTORC1) signaling, and thereby inhibition of autophagy and induction of endoplasmic reticulum stress in growth plate chondrocytes. IP injection of the mTORC1 inhibitor rapamycin to mice with Sirt1 deletion partially neutralized such inhibitory effects of Sirt1 ablation on longitudinal bone growth, indicating the causative link between SIRT1 and mTORC1 signaling in the growth plate. Mechanistically, SIRT1 interacted with tuberous sclerosis complex 2 (TSC2), a key upstream negative regulator of mTORC1 signaling, and loss of Sirt1 inhibited TSC2 expression, resulting in hyperactivated mTORC1 signaling in chondrocytes. In conclusion, our findings suggest that loss of Sirt1 may trigger mTORC1 signaling in growth plate chondrocytes and contributes to growth retardation, thus indicating that SIRT1 is an important regulator during chondrogenesis and providing new insights into the clinical potential of SIRT1 in bone development.

The role of the G protein-coupled receptor GPR30 in the effects of estrogen in ovariectomized mice

2009 ◽  
Vol 296 (3) ◽  
pp. E490-E496 ◽  
Author(s):  
S. H. Windahl ◽  
N. Andersson ◽  
A. S. Chagin ◽  
U. E. A. Mårtensson ◽  
H. Carlsten ◽  
...  
Keyword(s):  
Growth Plate ◽  
Fat Mass ◽  
Bone Growth ◽  
Distal Femur ◽  
Uterine Weight ◽  
Marrow Cellularity ◽  
G Protein Coupled ◽  
Estrogenic Responses

In vitro studies suggest that the membrane G protein-coupled receptor GPR30 is a functional estrogen receptor (ER). The aim of the present study was to determine the possible in vivo role of GPR30 as a functional ER primarily for the regulation of skeletal parameters, including bone mass and longitudinal bone growth, but also for some other well-known estrogen-regulated parameters, including uterine weight, thymus weight, and fat mass. Three-month-old ovariectomized (OVX) GPR30-deficient mice (GPR30−/−) and wild-type (WT) mice were treated with either vehicle or increasing doses of estradiol (E2; 0, 30, 70, 160, or 830 ng·mouse−1·day−1). Body composition [bone mineral density (BMD), fat mass, and lean mass] was analyzed by dual-energy-X ray absorptiometry, while the cortical and trabecular bone compartments were analyzed by peripheral quantitative computerized tomography. Quantitative histological analyses were performed in the distal femur growth plate. Bone marrow cellularity and distribution were analyzed using a fluorescence-activated cell sorter. The estrogenic responses on most of the investigated parameters, including increase in bone mass (total body BMD, spine BMD, trabecular BMD, and cortical bone thickness), increase in uterine weight, thymic atrophy, fat mass reduction, and increase in bone marrow cellularity, were similar for all of the investigated E2 doses in WT and GPR30−/− mice. On the other hand, E2 treatment reduced longitudinal bone growth, reflected by decreased femur length and distal femur growth plate height, in the WT mice but not in the GPR30−/− mice compared with vehicle-treated mice. These in vivo findings demonstrate that GPR30 is not required for normal estrogenic responses on several major well-known estrogen-regulated parameters. In contrast, GPR30 is required for a normal estrogenic response in the growth plate.

Sign in / Sign up

Export Citation Format

Share Document