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Plant villins: Versatile actin regulatory proteins

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Journal of Integrative Plant Biology 2015年第1期57卷 40-49页

作者： shanjin huang Xiaolu Qu Ruihui Zhang Key Laboratory of Plant Molecular Physiology Institute of Botany the Chinese Academy of Sciences

Regulation of actin dynamics is a central theme in cell biology that is important for different aspects of cell physiology. Villin, a member of the villin/gelsolin/fragmin superfamily of proteins, is an important regulator of actin. Villins contain six gelsolin homology domains （G1-G6） and an extra headpiece domain. In contrast to their mammalian counterparts, plant villins are expressed widely, implying that plant villins play a more general role in regulating actin dynamics. Some plant villins have a defined role in modifying actin dynamics in the pollentube; most of their in vivo activities remain to be ascertained. Recently, our understanding of the functions and mechanisms of action for plant villins has progressed rapidly, primarily due to the advent of Arabidopsis thaliana genetic approaches and imaging capabilities that can visualize actin dynamics at the single filament level in vitro and in living plant cells. In this review, we focus on discussing the biochemical activities and modes of regulation of plant villins. Here, we present current understand- ing of the functions of plant villins. Finally, we highlight some of the key unanswered questions regarding the functions and regulation of plant villins for future research.

关键词： Actin bundle actin capping actin cytoskeleton filamentsevering villin

Visualization of MicrotubuleOrganization and Dynamics in Living Arabidopsis Embryonic Cells

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Molecular Plant 2014年第8期7卷 1397-1401页

作者： Qiannan Wang shanjin huang Key Laboratory of Plant Molecular Physiology Institute of Botany Chinese Academy of Sciences Beijing 100093 China University of Chinese Academy of Sciences Beijing 100049 China National Center for Plant Gene Research Beijing 100101 China

Dear Editor, Embryogenesis is a critical developmental stage during the life cycle of flowering plants. During embryogenesis, the first round of asymmetric cell division in the zygote is followed by a series of cellular events, including cell division （symmetric or asymmetric） and directional cell expan- sion to generate the apical-basal axis, radial and lateral symmetry, and patterns of different cell fates for initiation of different organ primordia, which lay the foundation for post-embryonic development （Jurgens, 2001; Wendrich and Weijers, 2013）.

关键词： EMBRYOGENESIS CYTOSKELETON ENDOSPERM DIVISION PATTERN

Calcium imaging in Arabidopsis pollen cells using G-CaMP5

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Journal of Integrative Plant Biology 2018年第9期60卷 897-906页

作者： Min Diao Xiaolu Qu shanjin huang Key Laboratory of Plant Molecular Physiology Institute of BotanyChinese Academy of SciencesBeijing 100093China Center for Plant Biology School of Life SciencesTsinghua UniversityBeijing 100084China University of Chinese Academy of Sciences Beijing 100049China

Calcium （Ca2 ＋） signaling has been implicated in poJ]en germination and pollen tube growth. To date, however, we still know very little about how exactly Ca2＋ signaling links to various physiological subcellular processes during pollen germination and pollen tube growth. Given that Ca2＋ signaling is tightly related to the cytosolic concentration and dynamics of Ca2＋, it is vital to trace the dynamic changes in Ca＋ levels in order to decode Ca2＋ signaling. Here, we demonstrate that G-CaMP5 serves well as an indicator for monitoring cytosolic Ca2＋ dynamics in pollen cells. Using this probe, we show that cytosolic Ca2＋ changes dramatically during pollen germination, and, as reported previously, Ca2＋ forms a tip-focused gradient in the pollen tube and undergoes oscillation in the tip region during pollen tube growth. In particular, using G-CaMP5 allowed us to capture the dynamic changes in the cytosolic Ca2＋ concentration （[Ca2＋ ]cyt） in pollen tubes in response to various exogenous treatments. Our data suggest that G-CaMP5 is a suitable probe for monitoring the dynamics of [Ca2＋ ]cyt in pollen cells.

关键词： Calcium imaging in Arabidopsis pollen cells using G-CaMP5 MP

Regulation of Actin Dynamics in Pollen Tubes:Control of Actin Polymer Level

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Journal of Integrative Plant Biology 2009年第8期51卷 740-750页

作者： Naizhi Chen Xiaolu Qu Youjun Wu shanjin huang Center for Signal Transduction and Metabolomics Key Laboratory of Photosynthesis and Environmental Molecular Physiology Institute of Botany the Chinese Academy of Sciences Beijing 100093 China

Actin cytoskeleton undergoes rapid reorganization in response to internal and external cues. How the dynamics of actin cytoskeleton are regulated, and how its dynamics relate to its function are fundamental questions in plant cell biology. The pollen tube is a well characterized actin-based cell morphogenesis in plants. One of the striking features of actin cytoskeleton characterized in the pollen tube is its surprisingly low level of actin polymer. This special phenomenon might relate to the function of actin cytoskeleton in pollen tubes. Understanding the molecular mechanism underlying this special phenomenon requires careful analysis of actin-binding proteins that modulate actin dynamics directly. Recent biochemical and biophysical analyses of several highly conserved plant actin-binding proteins reveal unusual and unexpected properties, which emphasizes the importance of carefully analyzing their action mechanism and cellular activity. In this review, we highlight an actin monomer sequestering protein, a barbed end capping protein and an F-actin severing and dynamizing protein in plant. We propose that these proteins function in harmony to regulate actin dynamics and maintain the low level of actin polymer in pollen tubes.

关键词： actin-binding proteins actin dynamics pollen tube

In vitro assembly of plant tubulin in the absence of microtubule-stabilizing reagents

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Chinese Science Bulletin 2000年第24期45卷 2258-2263页

作者： huang shanjin REN Haiyun YUAN Ming Key Laboratory of Plant Physiology and Biochemistry of Ministry of Agriculture Department of Plant SciencesCollege of Biological SciencesChina Agricultural UniversityBeijing 100094China Key Laboratory of Cell Proliferation and Regulation Biology of Ministry of Education College of Life SciencesBeijing Normal UniversityBeijing 100875China

The assembly of microtubules is essential for physiological functions of microtubules. Addition of microtubule-stabilizing reagents or microtubule 'seeds' is usually necessary for plant tubulin assembly in vitro, which hinders the investigation of plant microtubule dynamics. In the present note, highly purified plant tubulins have been obtained from lily pollen, a non-microtubule-stabilizing reagent or microtubule 'seed' system for plant tubulin assembly has been established and the analysis of plant tubulin assembly performed. Experiment results showed that purified tubulin polymerized in vitro, and a typical microtubule structure was observed with electron microscopy. The kinetics curve of tubulin assembly exhibited typical 'parabola'. The presence of taxol significantly altered the character of plant tubulin assembly, including that abnormal microtubules were assembled and the critical concentration for plant tubulin assembly was decreased exceedingly from 3 mg/mL in the absence of taxol to 0.043 mg/

关键词： lily pollen tubulin microtubule tubulin assembly.

An Arabidopsis Class II Formin,AtFH19,Nucleates Actin Assembly,Binds to the Barbed End of Actin Filaments,and Antagonizes the Effect of AtFH1 on Actin Dynamics

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Journal of Integrative Plant Biology 2012年第10期54卷 800-813页

作者： Yiyan Zheng Haibo Xin Jinxing Lin Chun-Ming Liu shanjin huang Key Laboratory of Plant Molecular Physiology Institute of Botanythe Chinese Academy of SciencesBeijing 1%0093China University of Chinese Academy of Sciences Beijing 10049China Ornamental Horticulture and Landscape Architecture China Agricultural UniversityBeijing 100094China

Formin is a major protein responsible for regulating the nucleation of actin filaments, and as such, it permits the cell to control where and when to assemble actin arrays. It is encoded by a multigene family comprising 21 members in Arabidopsis thaliana. The Arabidopsis formins can be separated into two phylogenetically-distinct classes： there are 11 class I formins and 10 class II formins. Significant questions remain unanswered regarding the molecular mechanism of actin nucleation and elongation stimulated by each formin isovariant, and how the different isovariants coordinate to regulate actin dynamics in cells. Here, we characterize a class II formin, AtFH19, biochemically. We found that AtFH19 retains all general properties of the formin family, including nucleation and barbed end capping activity. It can also generate actin filaments from a pool of actin monomers bound to profilin. However, both the nucleation and barbed end capping activities of AtFH19 are less efficient compared to those of another well-characterized formin, AtFHI. Interestingly, AtFH19 FH1FH2 competes with AtFH1 FHIFH2 in binding actin filament barbed ends, and inhibits the effect of AtFH1 FHIFH2 on actin. We thus propose a mechanism in which two quantitatively different formins coordinate to regulate actin dynamics by competing for actin filament barbed ends.

关键词： Actin actin-binding proteins actin nucleation Arabidopsis cytoskeleton formin.

Organizational Innovation of Apical Actin Filaments Drives Rapid Pollen Tube Growth and Turning

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Molecular Plant 2017年第7期10卷 930-947页

作者： Xiaolu Qu Ruihui Zhang Meng Zhang Min Diao Yongbiao Xue shanjin huang Center for Plant Biology School of Life Sciences Tsinghua University Beijing 100084 China Key Laboratory of Plant Molecular Physiology Institute of Botany Chinese Academy of Sciences Beijing 100093 China State Key Laboratory of Molecular Developmental Biology Institute of Genetics and Developmental Biology Chinese Academy of Sciences Beijing 100101 China Beijing Institute of Genomics Chinese Academy of Sciences Beijing 100101 China

Polarized tip growth is a fundamental cellular process in many eukaryotes. In this study, we examined the dynamic restructuring of the actin cytoskeleton and its relationship to vesicle transport during pollen tip growth in Arabidopsis. We found that actin filaments originating from the apical membrane form a specialized structure consisting of longitudinally aligned actin bundles at the cortex and inner cytoplasmic fila- ments with a distinct distribution. Using actin-based pharmacological treatments and genetic mutants in combination with FRAP （fluorescence recovery after photobleaching） technology to visualize the transport of vesicles within the growth domain of pollen tubes, we demonstrated that cortical actin filaments facilitate tip-ward vesicle transport. We also discovered that the inner apical actin filaments prevent backward movement of vesicles, thus ensuring that sufficient vesicles accumulate at the pollen tube tip to support the rapid growth of the pollen tube. The combinatorial effect of cortical and internal apical actin filaments perfectly explains the generation of the inverted ＂V＂ cone-shaped vesicle distribution pattern at the pollen tube tip. When pollen tubes turn, apical actin filaments at the facing side undergo depolymerization and repolymerization to reorient the apical actin structure toward the new growth direction. This actin restructuring precedes vesicle accumulation and changes in tube morphology. Thus, our study provides new insights into the functional relationship between actin dynamics and vesicle transport during rapid and directional pollen tube growth.

关键词： pollen tube clear zone apical actin structure actin dynamics vesicle trafficking myosin

Profilin Regulates Apical Actin Polymerization to Control Polarized Pollen Tube Growth

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Molecular Plant 2015年第12期8卷 1694-1709页

作者： Xiaonan Liu Xiaolu Qu Yuxiang Jiang Ming Chang Ruihui Zhang Youjun Wu Ying Fu shanjin huang State Key Laboratory of Plant Physiology and Biochemistry College of Biological Sciences China Agricultural University Beijing 100193 China Key Laboratory of Plant Molecular Physiology Institute of Botany Chinese Academy of Sciences Beijing 100093 China Center for Plant Biology School of Life Sciences Tsinghua University Beijing 100084 China Tsinghua-Peking Joint Center for Life Sciences Beijing 100084 China National Center for Plant Gene Research Beijing 100101 China present address: Department of Biological Sciences Dartmouth College Hanover NH 03755 USA

Pollen tube growth is an essential step during flowering plant reproduction, whose growth depends on a population of dynamic apical actin filaments. Apical actin filaments were thought to be involved in the regu- lation of vesicle fusion and targeting in the pollen tube. However, the molecular mechanisms that regulate the construction of apical actin structures in the pollen tube remain largely unclear. Here, we identify profilin as an important player in the regulation of actin polymerization at the apical membrane in the pollen tube. Downregulation of profilin decreased the amount of filamentous actin and induced disorganization of apical actin filaments, and reduced tip-directed vesicle transport and accumulation in the pollen tube. Direct visualization of actin dynamics revealed that the elongation of actin filaments originating at the apical membrane decreased in profilin mutant pollen tubes. Mutant profilin that is defective in binding poly-L-proline only partially rescues the actin polymerization defect in profilin mutant pollen tubes, although it fully rescues the actin turnover phenotype. We propose that profilin controls the construction of actin structures at the pollen tube tip, presumably by favoring formin-mediated actin polymerization at the apical membrane.

关键词： pollen tube actin actin polymerization profilin formin

Molecular Mechanisms Underlying the Generation and Maintenance of Actin Cables in the Pollen Tube

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Molecular Mechanisms Underlying the Generation and Maintenan...