An inducible CiliaGFP mouse model for in vivo visualization (original) (raw)

An inducible CiliaGFP mouse model for in vivo visualization and analysis of cilia in live tissue

Cilia, 2013

Background: Cilia are found on nearly every cell type in the mammalian body, and have been historically classified as either motile or immotile. Motile cilia are important for fluid and cellular movement; however, the roles of non-motile or primary cilia in most tissues remain unknown. Several genetic syndromes, called the ciliopathies, are associated with defects in cilia structure or function and have a wide range of clinical presentations. Much of what we know about the formation and maintenance of cilia comes from model systems like C. elegans and Chalmydomonas. Studies of mammalian cilia in live tissues have been hampered by difficulty visualizing them.

Modeling ciliopathies: Primary cilia in development and disease

Current topics in developmental biology, 2008

Primary (nonmotile) cilia are currently enjoying a renaissance in light of novel ascribed functions ranging from mechanosensory to signal transduction. Their importance for key developmental pathways such as Sonic Hedgehog (Shh) and Wnt is beginning to emerge. The function of nodal cilia, for example, is vital for breaking early embryonic symmetry, Shh signaling is important for tissue morphogenesis and successful Wnt signaling for organ growth and differentiation. When ciliary function is perturbed, photoreceptors may die, kidney tubules develop cysts, limb digits multiply and brains form improperly. The etiology of several uncommon disorders has recently been associated with cilia dysfunction. The causative genes are often similar and their cognate proteins certainly share cellular locations and/or pathways. Animal models of ciliary gene ablation such as Ift88, Kif3a, and Bbs have been invaluable for understanding the broad function of the cilium. Herein, we describe the wealth of...

Various facets of vertebrate cilia: motility, signaling, and role in adult neurogenesis

Proceedings of The Japan Academy Series B-physical and Biological Sciences, 2009

Cilia are microtubule-based cellular organelles that are widely distributed in vertebrate tissues. They were rst observed hundreds of years ago. Recent studies indicate that this small organelle plays important roles in numerous physiological phenomena, including tissue morphogenesis, signal transduction, determination of left-right asymmetry during development, and adult neurogenesis. Ciliopathies, syndromes resulting from a genetic disorder of cilial components, frequently have complex eects involving many organ systems, owing to the broad distribution of cilia in the body.

The dynamic cilium in human diseases

PathoGenetics, 2009

Cilia are specialized organelles protruding from the cell surface of almost all mammalian cells. They consist of a basal body, composed of two centrioles, and a protruding body, named the axoneme. Although the basic structure of all cilia is the same, numerous differences emerge in different cell types, suggesting diverse functions. In recent years many studies have elucidated the function of 9+0 primary cilia. The primary cilium acts as an antenna for the cell, and several important pathways such as Hedgehog, Wnt and planar cell polarity (PCP) are transduced through it. Many studies on animal models have revealed that during embryogenesis the primary cilium has an essential role in defining the correct patterning of the body. Cilia are composed of hundreds of proteins and the impairment or dysfunction of one protein alone can cause complete loss of cilia or the formation of abnormal cilia. Mutations in ciliary proteins cause ciliopathies which can affect many organs at different levels of severity and are characterized by a wide spectrum of phenotypes. Ciliary proteins can be mutated in more than one ciliopathy, suggesting an interaction between proteins. To date, little is known about the role of primary cilia in adult life and it is tempting to speculate about their role in the maintenance of adult organs. The state of the art in primary cilia studies reveals a very intricate role. Analysis of cilia-related pathways and of the different clinical phenotypes of ciliopathies helps to shed light on the function of these sophisticated organelles. The aim of this review is to evaluate the recent advances in cilia function and the molecular mechanisms at the basis of their activity.

Biology of Cilia and Ciliopathies

Current Frontiers and Perspectives in Cell Biology, 2012

Biology of Cilia and Ciliopathies 425 These observations, however, are not entirely consistent among all ciliated organisms. Mutations in the kinesin-2 motor subunits of different species do not result in a cilia-less cell phenotype because of a secondary, homodimeric kinesin known as OSM-3 in C. elegans and KIF17 in Homo sapiens[13]. Studies investigating the function of OSM-3 conclude that the canonical kinesin-2 motor and OSM-3 work in a concerted effort to build sensory cilia in C. elegans [13]. Single mutants in KLP-11 (FLA8) and KAP-1 (FLA3) in C. elegans appear to form intact sensory cilia due to the redundancy of OSM-3 function in ciliogenesis (Signor et al, 1999). However, perturbations of OSM-3 results in loss of the ciliary distal segment comprised of singlet microtubule extensions beyond the doublet axoneme core. In these mutants, the heterotrimeric anterograde motor still allows formation of the middle segment. It could be possible the transferring of the IFT particle from the canonical kinesin-II to OSM-3 may insure proper, sequential construction of the cilia. However, it has been well documented that OSM-3 speed actually increases in disrupted kinesin-II mutants [21], suggesting that kinesin-II may in fact be negatively regulating OSM-3. If so, kinesin-II would ultimately be involved in determining the re-supply rate of axonemal precursors to the flagella compartment. Defects in retrograde IFT clearly demonstrate the negative impact that excess precursors and turnover products have on proper ciliary function. Therefore, accumulation of axonemal components, due to a faster influx of proteins by OSM-3, could also unbalance the natural turnover vs. assembly in favor of creating longer cilia, which is a phenotype that has been observed in kinesin-II mutants. Recently, a null mutant for a relatively new kinesin, KLP-6 in C. elegans males, demonstrated a slower procession of OSM-3/KAP-1-associated IFT particle within the ciliary compartment [22]. Although it was observed moving independently of the canonical IFT particle/motor complex, KLP-6 function may have a positive influence on ciliary length. This conclusion is supported by a reduction of klp-11/klp-6 double mutant cilia compared to the single klp-11 mutant; klp-11 mutant has comparatively longer cilia than wild-type. 2.2.1.2 Retrograde motor IFT-dynein, cytoplasmic dynein 2 (previously known as dynein 1b), powers the retrograde movement of IFT [12]. To date, four proteins are confirmed members of the dynein 2 complex: heavy chain DHC1b, light chain LC8, light intermediate chain D1bLIC, and an intermediate chain FAP133 [7-9, 14, 23-25]. C. elegans null mutants defective in dynein components undergo normal anterograde movement but accumulate large amounts of IFT proteins and turnover products within the ciliary compartment [26]. Retrograde-defective cilia are severely truncated and contain protein aggregates that appear as noticeable large, electron-dense clots. These results suggest IFT dynein is responsible for the retrograde movement of the IFT; this result has been seen in Chlamyomonas, where defects in IFT dynein lead to protein accumulations in the flagella compartment [27]. Anterograde movement remains active in these mutants; however, the characteristic bulbous cilia are present as a result of axonemal turnover outpacing the dysfunctional retrograde IFT. It has become fairly evident that IFT particles do not passively diffuse out of the flagella compartment and turnover products must be actively removed by dynein 2 in order to allow unhindered trafficking of the IFT trains. The current model for retrograde activation is fragmented at best. IFT-dynein is carried into the compartment in an inactivated form as part of the IFT cargo. Once it reaches the tip, a www.intechopen.com

The roles of cilia in developmental disorders and disease

Development, 2006

Cilia are highly conserved organelles that have diverse motility and sensory functions. Recent discoveries have revealed that cilia also have crucial roles in cell signaling pathways and in maintaining cellular homeostasis. As such, defects in cilia formation or function have profound effects on the development of body pattern and the physiology of multiple organ systems. By categorizing syndromes that are due to cilia dysfunction in humans and from studies in vertebrate model organisms, molecular pathways that intersect with cilia formation and function have come to light. Here, we summarize an emerging view that in order to understand some complex developmental pathways and disease etiologies, one must consider the molecular functions performed by cilia.

Identification of Cilia in Different Mouse Tissues

Cells

Cilia are microtubule-based hair-like organelles that extend from the cell surface. However, the existence and distribution of cilia in each organ and tissue at the postnatal stage in vivo remain largely unknown. In this study, we defined cilia distribution and arrangement and measured the ciliary lengths and the percentage of ciliated cells in different organs and tissues in vivo by using cilium dual reporter-expressing transgenic mice. Cilia were identified by the presence of ARL13B with an mCherry+ signal, and the cilium basal body was identified by the presence of Centrin2 with a GFP+ signal. Here, we provide in vivo evidence that chondrocytes and cells throughout bones have cilia. Most importantly, we reveal that: 1. primary cilia are present in hepatocytes; 2. no cilia but many centrioles are distributed on the apical cell surface in the gallbladder, intestine, and thyroid epithelia; 3. cilia on the cerebral cortex are well oriented, pointing to the center of the brain; 4. ARL...