scholarly journals A mouse model of BBS identifies developmental and homeostatic effects of BBS5 mutation and identifies novel pituitary abnormalities

2021 ◽  
Author(s):  
Melissa R Bentley-Ford ◽  
Staci E Engle ◽  
Kelsey R Clearman ◽  
Courtney J Haycraft ◽  
Reagan S Andersen ◽  
...  
Keyword(s):  
Cognitive Deficits ◽  
Primary Cilia ◽  
Protein Composition ◽  
Cell Types ◽  
Postnatal Life ◽  
Lacz Gene ◽  
Bardet Biedl Syndrome ◽  
Conditional Allele ◽  
Skeletal Defects ◽  
Ciliary Signaling

Abstract Primary cilia are critical sensory and signaling compartments present on most mammalian cell types. These specialized structures require a unique signaling protein composition relative to the rest of the cell to carry out their functions. Defects in ciliary structure and signaling result in a broad group of disorders collectively known as ciliopathies. One ciliopathy, Bardet–Biedl syndrome (BBS; OMIM 209900), presents with diverse clinical features, many of which are attributed to defects in ciliary signaling during both embryonic development and postnatal life. For example, patients exhibit obesity, polydactyly, hypogonadism, developmental delay and skeletal abnormalities along with sensory and cognitive deficits, but for many of these phenotypes it is uncertain, which are developmental in origin. A subset of BBS proteins assembles into the core BBSome complex, which is responsible for mediating transport of membrane proteins into and out of the cilium, establishing it as a sensory and signaling hub. Here, we describe two new mouse models for BBS resulting from a targeted LacZ gene trap allele (Bbs5−/−) that is a predicted congenital null mutation and conditional (Bbs5flox/flox) allele of Bbs5. Bbs5−/− mice develop a complex phenotype consisting of increased pre-weaning lethality craniofacial and skeletal defects, ventriculomegaly, infertility and pituitary anomalies. Utilizing the conditional allele, we show that the male fertility defects, ventriculomegaly and pituitary abnormalities are only present when Bbs5 is disrupted prior to postnatal day 7, indicating a developmental origin. In contrast, mutation of Bbs5 results in obesity, independent of the age of Bbs5 loss.

scholarly journals A Bbs5 mouse model reveals pituitary cilia contributions to developmental abnormalities

2020 ◽  
Author(s):  
Melissa R. Bentley ◽  
Staci E. Engle ◽  
Courtney J. Haycraft ◽  
Reagan S. Andersen ◽  
Mandy J. Croyle ◽  
...  
Keyword(s):  
Cognitive Deficits ◽  
Primary Cilia ◽  
Protein Composition ◽  
Cell Types ◽  
Postnatal Life ◽  
Bardet Biedl Syndrome ◽  
Developmental Abnormalities ◽  
Conditional Allele ◽  
Craniofacial Defects ◽  
Ciliary Signaling

AbstractPrimary cilia are critical sensory and signaling compartments present on most mammalian cell types. These specialized structures require a unique signaling protein composition relative to the rest of the cell to carry out their functions. Defects in ciliary structure and signaling result in a broad group of disorders collectively known as ciliopathies. One ciliopathy, Bardet-Biedl Syndrome (BBS; OMIM 209900), presents with diverse clinical features, many of which are attributed to defects in ciliary signaling during both embryonic development and postnatal life. For example, patients exhibit obesity, polydactyly, hypogonadism, developmental delay, and skeletal abnormalities along with sensory and cognitive deficits, but for many of these phenotypes it is uncertain which are developmental in origin. A subset of BBS proteins assembles into the BBSome complex, which is responsible for mediating transport of membrane proteins into and out of the cilium, establishing it as a sensory and signaling hub. Here we describe two new mouse models for BBS resulting from a congenital null and conditional allele of Bbs5. Bbs5 null mice develop a complex phenotype including craniofacial defects, skeletal shortening, ventriculomegaly, infertility, and pituitary anomalies. Utilizing the conditional allele, we show that the male fertility defects, ventriculomegaly, and pituitary abnormalities are only found when Bbs5 is mutated prior to P7 indicating a developmental origin. In contrast, mutation of Bbs5 results in obesity independent of the age of Bbs5 loss. Compared to other animal models of BBS, Bbs5 mutant mice exhibit pathologies that suggest a specialized role for Bbs5 in ciliary function.

scholarly journals Centrin2 regulates CP110 removal in primary cilium formation

2015 ◽  
Vol 208 (6) ◽  
pp. 693-701 ◽  
Author(s):  
Suzanna L. Prosser ◽  
Ciaran G. Morrison
Keyword(s):  
Calcium Binding ◽  
Primary Cilia ◽  
Reverse Genetics ◽  
Cell Types ◽  
Serum Starvation ◽  
Basal Bodies ◽  
Cilium Formation ◽  
Cellular Quiescence ◽  
Retinal Pigmented Epithelial Cells

Primary cilia are antenna-like sensory microtubule structures that extend from basal bodies, plasma membrane–docked mother centrioles. Cellular quiescence potentiates ciliogenesis, but the regulation of basal body formation is not fully understood. We used reverse genetics to test the role of the small calcium-binding protein, centrin2, in ciliogenesis. Primary cilia arise in most cell types but have not been described in lymphocytes. We show here that serum starvation of transformed, cultured B and T cells caused primary ciliogenesis. Efficient ciliogenesis in chicken DT40 B lymphocytes required centrin2. We disrupted CETN2 in human retinal pigmented epithelial cells, and despite having intact centrioles, they were unable to make cilia upon serum starvation, showing abnormal localization of distal appendage proteins and failing to remove the ciliation inhibitor CP110. Knockdown of CP110 rescued ciliation in CETN2-deficient cells. Thus, centrin2 regulates primary ciliogenesis through controlling CP110 levels.

scholarly journals Ciliary neuropeptidergic signaling dynamically regulates excitatory synapses in postnatal neocortical pyramidal neurons

2020 ◽  
Author(s):  
Lauren Tereshko ◽  
Ya Gao ◽  
Brian A. Cary ◽  
Gina G. Turrigiano ◽  
Piali Sengupta
Keyword(s):  
Primary Cilia ◽  
Neuronal Development ◽  
Pyramidal Neurons ◽  
Somatostatin Receptor ◽  
Cognitive Disorders ◽  
Mammalian Brain ◽  
Excitatory Synapses ◽  
Ciliary Signaling ◽  
Neocortical Pyramidal Neurons

ABSTRACTPrimary cilia are compartmentalized sensory organelles present on the majority of neurons in the mammalian brain throughout adulthood. Recent evidence suggests that cilia regulate multiple aspects of neuronal development, including the maintenance of neuronal connectivity. However, whether ciliary signals can dynamically modulate postnatal circuit excitability is unknown. Here we show that acute cell-autonomous knockdown of ciliary signaling rapidly strengthens glutamatergic inputs onto cultured neocortical pyramidal neurons, and increases spontaneous firing. This increased excitability occurs without changes to passive neuronal properties or intrinsic excitability. Further, the neuropeptide receptor somatostatin receptor 3 (SSTR3) is localized nearly exclusively to pyramidal neuron cilia both in vivo and in culture, and pharmacological manipulation of SSTR3 signaling bidirectionally modulates excitatory synaptic inputs onto these neurons. Our results indicate that ciliary neuropeptidergic signaling dynamically modulates excitatory synapses, and suggest that defects in this regulation may underlie a subset of behavioral and cognitive disorders associated with ciliopathies.

scholarly journals Genome Size and Structure Determine Efficiency of Postinternalization Steps and Gene Transfer of Capsid-Modified Adenovirus Vectors in a Cell-Type-Specific Manner

2004 ◽  
Vol 78 (18) ◽  
pp. 10009-10022 ◽  
Author(s):  
Dmitry M. Shayakhmetov ◽  
Zong-Yi Li ◽  
Anuj Gaggar ◽  
Helen Gharwan ◽  
Vladimir Ternovoi ◽  
...  
Keyword(s):  
Gene Transfer ◽  
Genome Size ◽  
First Generation ◽  
Viral Gene Expression ◽  
Protein Composition ◽  
Cell Types ◽  
Helper Virus ◽  
Viral Gene ◽  
Viral Genes ◽  
Size And Structure

ABSTRACT Adenovirus serotype 5 (Ad5) vectors containing Ad B-group fibers have become increasingly popular as gene transfer vectors because they efficiently transduce human cell types that are relatively refractory to Ad5 infection. So far, most B-group fiber-containing vectors have been first-generation vectors, deleted of E1 and/or E3 genes. Transduction with these vectors, however, results in viral gene expression and is associated with cytotoxicity and immune responses against transduced cells. To circumvent these problems, we developed fiber-chimeric Ad vectors devoid of all viral genes that were produced either by the homologous recombination of first-generation vectors or by using the Cre/lox-based helper virus system. In this study we compared early steps of infection between first-generation (35-kb genome) and Ad vectors devoid of all viral genes with genome sizes of 28 kb and 12.6 kb. All vectors possessed an Ad35-derived fiber knob domain, which uses CD46 as a primary attachment receptor. Using immortalized human hematopoietic cell lines and primary human CD34-positive hematopoietic cells, we found that the Ad genome size did not affect the efficiency of virus attachment to and internalization into cells. Furthermore, independently of the genome length and structure, all vectors migrated to the nucleus through late endosomal and lysosomal cellular compartments. However, the vector containing the short 12.6-kb genome was unable to efficiently escape from endosomes and deliver its DNA into the nucleus. Moreover, compared to other vectors, these Ad particles were less stable and had an abnormal capsid protein composition, including a lack of capsid-stabilizing protein IX. Our data indicate that the size and structure of the packaged viral genomes can affect the integrity of Ad particles, which in turn results in lower infectivity of Ad vectors.

Abstract 1433: The Cardioregulatory Peptide Apelin is Preferentially Expressed by Endothelial Cells and Upregulated in Myocardial Disease States

Circulation ◽  
2007 ◽  
Vol 116 (suppl_16) ◽  
Author(s):  
Ramendra K Kundu ◽  
Ahmad Y Sheikh ◽  
Michael Y Ho ◽  
Hyung J Chun ◽  
Diem T Huynh ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Reporter Gene ◽  
Fold Increase ◽  
Cell Types ◽  
Heart Tissue ◽  
Capillary Endothelium ◽  
Lacz Gene ◽  
Lacz Reporter ◽  
Rt Pcr ◽  
Disease States

Introduction: The APJ receptor and its ligand, apelin, comprise a homeostatic, cardio-regulatory pathway. Although cardiac apelin expression levels are altered in humans with cardiac failure, the cell type responsible for apelin expression and modulation in disease states remains unknown. Hypothesis: Apelin production is restricted to the endothelial compartment and is upregulated in states of cardiovascular stress. Methods: Transgenic apelin-LacZ reporter mice (SVJ background) were created by insertion of the nuclear localizing bacterial LacZ gene immediately downstream of the apelin promoter. Mice (n=12) were randomized to left anterior descending coronary artery (LAD) ligation, thoracic aortic constriction (TAC) or sham groups. Hearts were harvested 3 and 8 weeks post-TAC or LAD ligation, respectively. Localization of apelin expression was determined by Xgal staining. Endothelial phenotype of lacZ positive cells was confirmed by CD31 co-staining. Apelin expressing cells were quantified by histology. Apelin reporter results were confirmed by quantitating apelin expression in WT animals following LAD ligation (n=11) or sham (n=11) procedure by RT-PCR. Results: Extensive immunohistochemistry studies of heart tissue revealed lacZ reporter gene expression to be restricted to the coronary venous and capillary endothelium, with no expression by cardiomyocytes. Following both LAD ligation and TAC, the number of LacZ-apelin (+) endothelial cells significantly increased (p<0.002) in all chambers of the heart (Table ), with no evidence of apelin expression by other cell types. Evaluation of WT hearts by RT-PCR for the apelin gene confirmed the reporter gene findings with 1.3±0.3 fold increase (p<0.05) of apelin expression induced by LAD ligation compared to sham. Conclusions: Apelin is primary expressed by endothelial cells within the heart and is upregulated in response to myocardial stress. Apelin-LacZ Expressing Endothelial Cells are Increased Following Myocardial Injury

scholarly journals The N-myc proto-oncogene and IGF-II growth factor mRNAs are expressed by distinct cells in human fetal kidney and brain.

1989 ◽  
Vol 108 (3) ◽  
pp. 1093-1104 ◽  
Author(s):  
H Hirvonen ◽  
M Sandberg ◽  
H Kalimo ◽  
V Hukkanen ◽  
E Vuorio ◽  
...  
Keyword(s):  
Growth Factor ◽  
Wilms Tumor ◽  
Expression Patterns ◽  
Regional Distribution ◽  
Cell Types ◽  
Cortical Plate ◽  
Northern Hybridization ◽  
Postnatal Life ◽  
Fetal Kidney ◽  
The Brain

We studied the expression of the N-myc proto-oncogene and the insulin-like growth factor-II (IGF-II) gene in human fetuses of 16-19 gestational wk. Both genes have specific roles in the growth and differentiation of embryonic tissues, such as the kidney and neural tissue. Since continued expression of N-myc and IGF-II mRNAs is also a characteristic feature of Wilms' tumor, a childhood neoplasm of probable fetal kidney origin, we were particularly interested in the possibility that their expression might be linked or coordinately regulated in the developing kidney. Expression of N-myc mRNA was observed in the brain and in the kidney by Northern hybridization analysis. In in situ hybridization of the kidney, N-myc autoradiographic grains were primarily located over epithelially differentiating mesenchyme while most of the mesenchymal stromal cells showed only a background signal with the N-myc probe. N-myc mRNA was detectable throughout the developing brain with a slight accentuation in the intermediate zone cells in between the subependymal and cortical layers. Thus, even postmitotic neuroepithelial cells of the fetal cerebrum expressed N-myc mRNA. In Northern hybridization, IGF-II mRNA signal was abundant in the kidney but much weaker, though definite, in the brain. The regional distribution of IGF-II mRNA in the kidney was largely complementary to that of N-myc. IGF-II autoradiographic grains were located predominantly over the stromal and blastemal cells with a relative lack of hybridization over the epithelial structures. In the brain, IGF-II mRNA was about two- to threefold more abundant in the subependymal and intermediate layers than in the cortical plate and ependymal zone, respectively. The fetal expression patterns of the N-myc and IGF-II mRNAs are reflected by the types of tumors known to express the corresponding genes during postnatal life such as Wilms' tumor. However, the apparent coexpression of the IGF-II and N-myc genes in immature kidneys occurs largely in distinct cell types.

scholarly journals Primary Cilia and Calcium Signaling Interactions

2020 ◽  
Vol 21 (19) ◽  
pp. 7109
Author(s):  
Hannah Saternos ◽  
Sidney Ley ◽  
Wissam AbouAlaiwi
Keyword(s):  
Primary Cilia ◽  
Cell Types ◽  
Cellular Level ◽  
Cellular Functions ◽  
Complex Processes ◽  
Secondary Messenger ◽  
Body Patterning ◽  
Motile Cilia ◽  
And Migration ◽  
Context Specific

The calcium ion (Ca2+) is a diverse secondary messenger with a near-ubiquitous role in a vast array of cellular processes. Cilia are present on nearly every cell type in either a motile or non-motile form; motile cilia generate fluid flow needed for a variety of biological processes, such as left–right body patterning during development, while non-motile cilia serve as the signaling powerhouses of the cell, with vital singling receptors localized to their ciliary membranes. Much of the research currently available on Ca2+-dependent cellular actions and primary cilia are tissue-specific processes. However, basic stimuli-sensing pathways, such as mechanosensation, chemosensation, and electrical sensation (electrosensation), are complex processes entangled in many intersecting pathways; an overview of proposed functions involving cilia and Ca2+ interplay will be briefly summarized here. Next, we will focus on summarizing the evidence for their interactions in basic cellular activities, including the cell cycle, cell polarity and migration, neuronal pattering, glucose-mediated insulin secretion, biliary regulation, and bone formation. Literature investigating the role of cilia and Ca2+-dependent processes at a single-cellular level appears to be scarce, though overlapping signaling pathways imply that cilia and Ca2+ interact with each other on this level in widespread and varied ways on a perpetual basis. Vastly different cellular functions across many different cell types depend on context-specific Ca2+ and cilia interactions to trigger the correct physiological responses, and abnormalities in these interactions, whether at the tissue or the single-cell level, can result in diseases known as ciliopathies; due to their clinical relevance, pathological alterations of cilia function and Ca2+ signaling will also be briefly touched upon throughout this review.

scholarly journals Reproduction Function in Male Patients With Bardet Biedl Syndrome

2020 ◽  
Vol 105 (12) ◽  
pp. e4417-e4429
Author(s):  
Isabelle Koscinski ◽  
Manuel Mark ◽  
Nadia Messaddeq ◽  
Jean Jacques Braun ◽  
Catherine Celebi ◽  
...  
Keyword(s):  
Primary Cilia ◽  
Wide Spectrum ◽  
Testis Size ◽  
Sexual Hormones ◽  
Bardet Biedl Syndrome ◽  
Self Confidence ◽  
Electron Microscopy Analysis ◽  
Microscopy Analysis ◽  
Male Patients ◽  
Child Wish

Abstract Purpose Bardet-Biedl syndrome (BBS) is a ciliopathy with a wide spectrum of symptoms due to primary cilia dysfunction, including genitourinary developmental anomalies as well as impaired reproduction, particularly in males. Primary cilia are known to be required at the following steps of reproduction function: (i) genitourinary organogenesis, (ii) in fetal firing of hypothalamo-pituitary axe, (iii) sperm flagellum structure, and (iv) first zygotic mitosis conducted by proximal sperm centriole. BBS phenotype is not fully understood. Methods This study explored all steps of reproduction in 11 French male patients with identified BBS mutations. Results BBS patients frequently presented with genitourinary malformations, such as cryptorchidism (5/11), short scrotum (5/8), and micropenis (5/8), but unexpectedly, with normal testis size (7/8). Ultrasonography highlighted epididymal cysts or agenesis of one seminal vesicle in some cases. Sexual hormones levels were normal in all patients except one. Sperm numeration was normal in 8 out of the 10 obtained samples. Five to 45% of sperm presented a progressive motility. Electron microscopy analysis of spermatozoa did not reveal any homogeneous abnormality. Moreover, a psychological approach pointed to a decreased self-confidence linked to blindness and obesity explaining why so few BBS patients express a child wish. Conclusions Primary cilia dysfunction in BBS impacts the embryology of the male genital tract, especially epididymis, penis, and scrotum through an insufficient fetal androgen production. However, in adults, sperm structure does not seem to be impacted. These results should be confirmed in a greater BBS patient cohort, focusing on fertility.

Antigenic variation and increase in pathogenicity in Pseudomonas aeruginosa as a result of growth on glucose or N-hexadecane as sole carbon source

1978 ◽  
Vol 24 (6) ◽  
pp. 675-679 ◽  
Author(s):  
Robert S. Stinson ◽  
D. E. Talburt
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Carbon Source ◽  
Outer Membrane ◽  
Sole Carbon Source ◽  
Antigenic Variation ◽  
Protein Composition ◽  
Cell Types ◽  
Additional Protein

When Pseudomonas aeruginosa is grown on glucose as opposed to n-hexadecane as the sole carbon source, the antigenicity, virulence, and protein composition of the outer membrane are altered. The hydrocarbon-grown cells demonstrate a 3-log increase in virulence over the glucose-grown cells (in mice). There also appears to be an additional protein present in the outer membrane of the n-hexadecane-grown cells. This protein may contribute to the observed antigenic differences between the two cell types.

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