Chapter 17 - Regulation of Cilia assembly, Disassembly, and Length by Protein Phosphorylation

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Abstract

The exact mechanism by which cells are able to assemble, regulate, and disassemble cilia or flagella is not yet completely understood. Recent studies in several model systems, including Chlamydomonas, Tetrahymena, Leishmania, Caenorhabditis elegans, and mammals, provide increasing biochemical and genetic evidence that phosphorylation of multiple protein kinases plays a key role in cilia assembly, disassembly, and length regulation. Members of several protein kinase families—including aurora kinases, never in mitosis A (NIMA)-related protein kinases, mitogen-activated protein (MAP) kinases, and a novel cyclin-dependent protein kinase—are involved in the ciliary regulation process. Among the newly identified protein kinase substrates are Chlamydomonas kinesin-13 (CrKinesin13), a microtubule depolymerizer, and histone deacetylase 6 (HDAC6), a microtubule deacetylase. Chlamydomonas aurora/Ipl1p-like protein kinase (CALK) and CrKinesin13 are two proteins that undergo phosphorylation changes correlated with flagellar assembly or disassembly. CALK becomes phosphorylated when flagella are lost, whereas CrKinesin13 is phosphorylated when new flagella are assembled. Conversely, suppressing CrKinesin13 expression results in cells with shorter flagella.

Introduction

The microtubule-based cellular organelles, cilia and eukaryotic flagella, serve as motors and cellular antennae (Fliegauf et al., 2007, Gerdes et al., 2009, Pan, 2008). They are ancient organelles, well conserved throughout the biological kingdom, and present in organisms ranging from protists to mammals. For historical reasons, cilia and eukaryotic flagella actually refer to the same cellular organelle; thus, these two terms are used interchangeably.

A cilium is anchored to the cell body through a structure called the basal body, which is derived from the centriole. The length of a cilium is cell-type-specific and is maintained during G1 or G0 phase of the cell cycle so that the cilium can properly perform its function. Cilia may be lost in response to environmental stress or development signals by two distinct mechanisms: gradually shortening from the cilia tip (resorption) or detachment from the cilia base (deflagellation). When cilia are lost, cells are able to regenerate new cilia of the same length. Cilia are resorbed before mitosis, thereby releasing the centrioles to participate in spindle formation. After cytokinesis, cilia are formed on the centrioles. In vertebrates, a primary cilium is typically formed on the mother centriole in uniciliated cells. In the eukaryotic organism Chlamydomonas, however, two flagella are assembled by nucleating microtubules from both the mother and daughter centrioles. Cilia formation in multiciliated cells involves the generation of multiple centrioles either spontaneously (acentriolar pathway) or through duplication of existing centrioles (centriolar pathway) (Hagiwara et al., 2000, Sorokin, 1962, Sorokin, 1968). The assembly, length regulation, and loss of cilia are complex cellular processes involving gene expression, protein transport, remodeling of protein complexes, and signal transduction.

Although cilia structure, function, and consequences of their defects are largely known, less is understood about the regulatory network involved in cilia assembly, disassembly, and length maintenance. Protein phosphorylation has emerged as a key mechanism for regulating these biological processes. We will discuss recent evidence on the function of protein phosphorylation in the control of each step of the cilia “life cycle”: assembly, disassembly, and length control.

Section snippets

Protein Phosphorylation in Cilia Assembly

Cilia formation requires multiple cellular processes: signaling to initiate cilia assembly, expression of ciliary genes, remodeling of cytoplasmic microtubules, protein transport to the cilia assembly site, and posttranslational modification of axonemal microtubules. Exciting discoveries have recently been made regarding phosphorylation control of cilia assembly, which will open new research opportunities for studying ciliary regulation.

Protein Phosphorylation in Cilia Disassembly

Before mitosis, primary cilia are lost in order to release the basal body to engage in mitosis (Pan and Snell, 2007, Quarmby and Parker, 2005). The precise phase of the cell cycle during which cilia are lost has not been definitively determined; different researchers have proposed different answers, from the S phase, to the G2/M phase, to prophase (Pugacheva et al., 2007, Rieder et al., 1979, Tucker et al., 1979). Cells may lose cilia by deflagellation or cilia resorption. The current evidence

Protein Phosphorylation in Cilia Length Control

Members of divergent protein kinase families play a role in controlling cilia length (Wilson et al., 2008). The ciliary function of these protein kinases has largely been revealed by mutant analysis, RNAi suppression, and/or gene overexpression.

Conclusion

The protein kinases and/or their substrates that have been implicated in ciliary regulation are depicted in Fig. 1. The phosphoproteins involved in assembly and/or disassembly of cilia (e.g., LF2p) may control ciliary length as well. However, phosphoproteins that regulate flagellar length may not be required for flagellar assembly (e.g., LF4p). Genetic evidence supports the idea that most of these phosphoproteins play a role in ciliary regulation; however, relevant biochemical evidence or

Acknowledgments

We are indebted to Dr. William Snell from UT Southwestern Medical Center, Dallas, for insightful suggestion and comments on the manuscript. This work was supported in part by the National Natural Science Foundation of China (Nos. 30671090, 30830057, and 30771084) and the National Basic Research Program of China (No. 2007CB914401) (to JP).

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