拟南芥叶绿体分裂蛋白PARC6影响子叶与真叶的生长

doi:10.7525/j.issn.1673-5102.2023.05.007

摘要/Abstract

摘要：

在DNA、RNA及细胞水平上鉴定了叶绿体异常分裂纯合突变株parc6，子叶白化纯合突变株sco2，同时以真叶异常分裂及子叶白化双重突变株sl2作为参照，通过向培养基中施加不同浓度的蔗糖含量来探究叶绿体异常分裂对子叶和真叶生长的影响。结果显示，sco2突变株子叶白化，而真叶生长正常；parc6突变株子叶生长势及生活力显著低于野生型对照，相当于sco2突变株，而子叶白化与真叶异常分裂双重突变株sl2生长严重受阻。parc6突变株真叶的生长势与生活力也显著低于野生型，但相较于子叶则有所恢复。parc6突变株子叶生长受阻表型可以通过在培养基上补充碳源而得到恢复，sco2突变株子叶生长受阻表型可以通过补充碳源得到恢复，但叶绿体荧光参数相较于野生型仍有差异。蓝色温和凝胶电泳结合二向聚丙烯酰胺凝胶电泳结果表明parc6突变株的子叶及真叶光系统高级结构组装正常，说明parc6异常分裂的叶绿体无法生成足够的能量进而影响子叶及真叶的生长。进化分析表明，PARC6与SCO2存在共同进化的趋势，说明子叶发育与叶绿体分裂存在一定的联系。结果指示，叶绿体大小与植物生长尤其是子叶的发育存在密切的联系，这为深入揭示叶绿体的功能提供一个新的视角。

关键词: 拟南芥, 叶绿体分裂, PARC6, 子叶, 真叶

Abstract:

Abnormal chloroplast division homozygous mutant strain parc6 and albinistic cotyledon homozygous mutant strain sco2 were identified at the DNA， RNA and cellular levels， while the effects of chloroplast abnormal division on the growth of cotyledon and leaf were investigated by adding different concentrations of sucrose to the medium and taking sl2， the double mutant with leaf abnormal division and cotyledon albino as the control. The results showed that the cotyledon of sco2 mutant was albino， while the leaf grew normally. Meanwhile， the cotyledon growth and viability of the parc6 mutant were significantly lower than the wild-type， which was equivalent to the sco2 mutant， however， the double mutant sl2 with cotyledon albinism and leaf abnormal division was severely hindered. The leaf growth and viability of the parc6 mutant were also significantly lower than that of the wild type， but recovered compared to the cotyledon. The inhibited cotyledon growth phenotype of the parc6 mutant and sco2 mutant could be confirmed by the addition of carbon source to the medium， but chloroplast fluorescence parameters of sco2 were different from the wild type. The results of BN-PAGE combined with SDS-PAGE showed that the high-level structure of the photosystem of the cotyledon and leaf were normal， indicating that chloroplasts of parc6 failed to produce enough energy to affect the growth of cotyledon and leaf. The phylogenetic analysis showed that PARC6 and SCO2 coevolved， suggesting a connection between cotyledon development and chloroplast division. Our results indicated that chloroplast size was closely related to plant growth， especially the development of cotyledon， which provided a new perspective for revealing chloroplast function.

Key words: Arabidopsis thaliana, Chloroplast division, PARC6, Cotyledons, leaf

中图分类号:

Q751

江转转, 龚莉, 宋亚玲. 拟南芥叶绿体分裂蛋白PARC6影响子叶与真叶的生长[J]. 植物研究, 2023, 43(5): 700-710.

Zhuanzhuan JIANG, Li GONG, Yaling SONG. The Chloroplast Division Protein PARC6 Affected the Growth of Cotyledon and Leaf in Arabidopsis thaliana[J]. Bulletin of Botanical Research, 2023, 43(5): 700-710.

图/表 9

表1

图1

图2

图3

图4

图5

图6

图7

图8

参考文献 35

1	RAVEN J A， ALLEN J F.Genomics and chloroplast evolution： what did cyanobacteria do for plants？［J］.Genome Biology，2003，4（3）：209.
2	POGSON B J， GANGULY D， ALBRECHT-BORTH V.Insights into chloroplast biogenesis and development［J］.Biochimica et Biophysica Acta（BBA）-Bioenergetics，2015，1847（9）：1017-1024.
3	SUMIYA N， FUJIWARA T，ERA A，et al.Chloroplast division checkpoint in eukaryotic algae［J］.Proceedings of the National Academy of Sciences of the United States of America，2016，113（47）：E7629-E7638.
4	MIYAGISHIMA S Y.Mechanism of plastid division：from a bacterium to an organelle［J］.Plant Physiology，2011，155（4）：1533-1544.
5	CHEN L， SUN B， GAO W，et al.MCD1 associates with FtsZ filaments via the membrane-tethering protein ARC6 to guide chloroplast division［J］.The Plant Cell，2018，30（8）：1807-1823.
6	ZHANG Y H， ZHANG X C， CUI H S，et al.Residue 49 of AtMinD1 plays a key role in the guidance of chloroplast division by regulating the ARC6-AtMinD1 Interaction［J］.Frontiers in Plant Science，2021，12：1-16.
7	PORTER K J， CAO L Y， CHEN Y D，et al.The Arabidopsis thaliana chloroplast division protein FtsZ1 counterbalances FtsZ2 filament stability in vitro ［J］.The Journal of Biological Chemistry，2021，296：100627.
8	SUN B， ZHANG Q Y， YUAN H，et al.PDV1 and PDV2 differentially affect remodeling and assembly of the chloroplast DRP5B ring［J］.Plant Physiology，2020，182（4）：1966-1978.
9	OHASHI Y， MORI T， IGAWA T.Behavior of filamentous temperature-sensitive Z2 （FtsZ2） in the male gametophyte during sexual reproduction processes of flowering plants［J］.Protoplasma，2020，257（4）：1201-1210.
10	MIYAGISHIMA S Y， NISHIDA K， MORI T，et al.A plant-specific dynamin-related protein forms a ring at the chloroplast division site［J］.The Plant Cell，2003，15（3）：655-665.
11	OKAZAKI K， KABEYA Y， MIYAGISHIMA S Y.The evolution of the regulatory mechanism of chloroplast division［J］.Plant Signaling & Behavior，2010，5（2）：164-167.
12	ZHANG M， HU Y， JIA J J，et al.CDP1，a novel component of chloroplast division site positioning system in Arabidopsis ［J］.Cell Research，2009，19（7）：877-886.
13	GLYNN J M， YANG Y， VITHA S，et al.PARC6，a novel chloroplast division factor，influences FtsZ assembly and is required for recruitment of PDV1 during chloroplast division in Arabidopsis ［J］.The Plant Journal，2009，59（5）：700-711.
14	ISHIKAWA H， YASUZAWA M， KOIKE N，et al. Arabidopsis PARC6 is critical for plastid morphogenesis in pavement，trichome，and guard cells in leaf epidermis［J］.Frontiers in Plant Science，2020，10：1665.
15	CHEN C， CAO L Y， YANG Y，et al.ARC3 activation by PARC6 promotes FtsZ-ring remodeling at the chloroplast division site［J］.The Plant Cell，2019，31（4）：862-885.
16	MECHELA A， SCHWENKERT S， SOLL J.A brief history of thylakoid biogenesis［J］.Open Biology，2019，9（1）：180237.
17	BÖRNER T， ALEYNIKOVA A Y， ZUBO Y O，et al.Chloroplast RNA polymerases：role in chloroplast biogenesis［J］.Biochimica et Biophysica Acta （BBA） - Bioenergetics，2015，1847（9）：761-769.
18	WANG Y N， JIE W G， PENG X Y，et al.Physiological adaptive strategies of oil seed crop ricinus communis early seedlings（cotyledon vs.true leaf） under salt and alkali stresses：from the growth，photosynthesis and chlorophyll fluorescence［J］.Frontiers in Plant Science，2019，9：1939.
19	ISHIZAKI Y， TSUNOYAMA Y， HATANO K，et al.A nuclear-encoded sigma factor，Arabidopsis SIG6，recognizes sigma-70 type chloroplast promoters and regulates early chloroplast development in cotyledons［J］.The Plant Journal，2005，42（2）：133-144.
20	TANZ S K， KILIAN J， JOHNSSON C，et al.The SCO2 protein disulphide isomerase is required for thylakoid biogenesis and interacts with LCHB1 chlorophyll a/b binding proteins which affects chlorophyll biosynthesis in Arabidopsis seedlings［J］.The Plant Journal，2012，69（5）：743-754.
21	ALBRECHT V， INGENFELD A， APEL K.Snowy cotyledon 2：the identification of a zinc finger domain protein essential for chloroplast development in cotyledons but not in true leaves［J］.Plant Molecular Biology，2008，66（6）：599-608.
22	QI Y F， WANG X M， LEI P，et al.The chloroplast metalloproteases VAR2 and EGY1 act synergistically to regulate chloroplast development in Arabidopsis ［J］.The Journal of Biological Chemistry，2020，295（4）：1036-1046.
23	QI Y F， LIU X Y， LIANG S，et al.A putative chloroplast thylakoid metalloprotease VIRESCENT3 regulates chloroplast development in Arabidopsis thaliana ［J］.The Journal of Biological Chemistry，2016，291（7）：3319-3332.
24	JIANG Z Z， ZHU L， WANG Q Y，et al.Autophagy-related 2 regulates chlorophyll degradation under abiotic stress conditions in Arabidopsis ［J］.International Journal of Molecular Sciences，2020，21（12）：4515.
25	FU A G， HE Z Y， CHO H S，et al.A chloroplast cyclophilin functions in the assembly and maintenance of photosystem Ⅱ in Arabidopsis thaliana ［J］.Proceedings of the National Academy of Sciences of the United States of America，2007，104（40）：15947-15952.
26	TAMURA K， STECHER G， KUMAR S.MEGA11：molecular evolutionary genetics analysis version 11［J］.Molecular Biology and Evolution，2021，38（7）：3022-3027.
27	COPE A L， O'MEARA B C， GILCHRIST M A.Gene expression of functionally-related genes coevolves across fungal species：detecting coevolution of gene expression using phylogenetic comparative methods［J］.BMC Genomics，2020，21（1）：370.
28	METEIGNIER L V， GHANDOUR R， MEIERHOFF K，et al.The Arabidopsis mTERF-repeat MDA1 protein plays a dual function in transcription and stabilization of specific chloroplast transcripts within the psbE and ndhH operons［J］.New Phytologist，2020，227（5）：1376-1391.
29	HE Y， SHI Y F， ZHANG X B，et al.The OsABCI7 transporter interacts with OsHCF222 to stabilize the thylakoid membrane in Rice［J］.Plant Physiology，2020，184（1）：283-299.
30	DUTTA S， CRUZ J A， IMRAN S M，et al.Variations in chloroplast movement and chlorophyll fluorescence among chloroplast division mutants under light stress［J］.Journal of Experimental Botany，2017，68（13）：3541-3555.
31	YOSHIDA Y.Insights into the mechanisms of chloroplast division［J］.International Journal of Molecular Sciences，2018，19（3）：733.
32	PIPITONE R， EICKE S， PFISTER B，et al.A multifaceted analysis reveals two distinct phases of chloroplast biogenesis during de-etiolation in Arabidopsis ［J］.eLife，2021，10：e62709.
33	SUN T H， ZHOU F， HUANG X Q，et al.Orange represses chloroplast biogenesis in etiolated Arabidopsis cotyledons via interaction with TCP14［J］.The Plant Cell，2019，31（12）：2996-3014.
34	HSIEH W Y， SUNG T Y， WANG H T，et al.Functional evidence for the critical amino-terminal conserved domain and key amino acids of Arabidopsis 4-Hydroxy-3-methylbut-2-enyl diphosphate reductase［J］.Plant Physiology，2014，166（1）：57-69.
35	WU G Z， XUE H W. Arabidopsis β-ketoacyl-［acyl carrier protein］ synthase i is crucial for fatty acid synthesis and plays a role in chloroplast division and embryo development［J］.The Plant Cell，2010，22（11）：3726-3744.

基因名称 Gene name		引物序列5′→3′ Primer sequence 5′→3′
PCR引物 PCR primer	SCO2	F：CTTCTTCCTCCATTTTCGTTACACCT
	SCO2	R：GTCCATTGTTGAGTTTCACATT
	PARC6	LP：TCTCGCACATTAGTTATGGGC
	PARC6	RP：ATAGGTCTTAACCTGGCGAGC
	PARC6	TP：ATTTTGCCGATTTCGGAAC
QPCR引物 QPCR primer	SCO2	F：AATGGCTTCTTCTTTCACCAATAGTC
	SCO2	R：GGTGTCTCGAGGTCTGATACATATAAAA
	PARC6	F：GTCCTTGATGAATCCATGCTTGTCCAG
	PARC6	R：GAAGAGCTTCGATTTCTGCAGCCTCACC
	ACTIN2	F：CCCGATGGGCAAGTCATC
	ACTIN2	R：GAACAAGACTTCTGGGCATCTGA

[1]	郑晟, 高海霞, 苏敏, 卢尚欢, 张腾国, 武国凡. 外源蔗糖影响AtKEA1和AtKEA2调节拟南芥幼苗根的生长[J]. 植物研究, 2023, 43(4): 562-571.
[2]	裘喻平, 王益川, 郭红卫. 植物根毛发育调控机制的研究进展[J]. 植物研究, 2023, 43(3): 321-332.
[3]	蔡圆圆, 夏季奔奔, 应文涵, 王洁瑶, 谢涛, 邢孔丫, 冯宣军, 华学军. 拟南芥线粒体蛋白突变体ssr1-2表型的详细鉴定与分析[J]. 植物研究, 2023, 43(3): 421-431.
[4]	王梦姣, 曹钰雪, 徐永盛, 丁风鹅, 苏乔. 过表达海洋微生物宏基因组MbCSP提高转基因拟南芥的抗旱和耐寒性[J]. 植物研究, 2022, 42(2): 243-251.
[5]	田红梅, 郭霄, 王奎玲, 孙迎坤. 耐冬山茶子叶再生体系的研究[J]. 植物研究, 2021, 41(3): 321-328.
[6]	武国凡, 成宏斌, 吴玉俊, 沈娟, 吴旺泽. CRISPR/Cas9介导靶向敲除拟南芥BRI1突变体的鉴定[J]. 植物研究, 2021, 41(3): 362-371.
[7]	张雨晴, 刘野, 曲春浦, 刘关君, 杨天天, 杨成君. 转基因PnDof30拟南芥非生物胁迫下的抗性分析[J]. 植物研究, 2020, 40(3): 407-415.
[8]	李子义, 贺子航, 卢惠君, 王玉成, 及晓宇. 拟南芥AtUNE12基因的耐盐功能初探[J]. 植物研究, 2020, 40(2): 257-265.
[9]	何好, 朱国庆, 陈诗雅, 徐畅, 金淑梅. 细叶百合LpPEX7基因克隆及盐胁迫下的表达特性分析[J]. 植物研究, 2020, 40(2): 274-283.
[10]	王爽, 程玉祥, 夏德安. 拟南芥根毛功能基因AtGDPD-Like3关键氨基酸位点鉴定[J]. 植物研究, 2020, 40(1): 79-84.
[11]	李芃, 郇兆蔚, 丁兰. Rabdosinate调节生长素极性运输蛋白PIN1、PIN3和PIN4抑制拟南芥幼苗根生长[J]. 植物研究, 2019, 39(6): 908-916.
[12]	李爽, 熊樱, RALF M;ller-Xing, 邢倩. 转录因子WRKY6和PR1在拟南芥胁迫记忆中的表达模式[J]. 植物研究, 2019, 39(5): 752-759.
[13]	樊二勤, 刘彩霞, 付鹏跃, 杨传平, 曲冠证. 84K杨PagC3H3基因表达模式分析[J]. 植物研究, 2019, 39(4): 521-528.
[14]	林云, 毕海燕, 李超, 云映霞. 中国11个双子叶植物原白中排印错误之更正[J]. 植物研究, 2019, 39(2): 310-320.
[15]	赵敏, 王玥萱, 徐运飞, 赵启安, 刘博, 杨宁. 干旱胁迫下拟南芥中H₂S与ABA信号关系研究[J]. 植物研究, 2019, 39(1): 104-112.