Identification of CaLOP Regulating Development and Growth Through Virus-Induced Gene Silencing in Pepper

Won-Hee   Kang; Junesung  Lee; Jae-Young  Nam; Seon-In  Yeom

doi:10.12972/kjhst.20180029

All Issue

2018 Vol.36, Issue 2 Preview Page Next Page

Identification of CaLOP Regulating Development and Growth Through Virus-Induced Gene Silencing in Pepper 유전자 침묵현상을 이용한 고추 리포옥시게네이즈 유사 유전자의 생육과 발달 관련 기능 분석

30 April 2018. pp. 292-302

PDF XML

Abstract

The proteins secreted into the cell wall and extracellular space have multiple roles in plant development, metabolism, and stress responses. In a previous study, examples of such proteins secreted during pepper–pathogen interactions were isolated to elucidate their roles. Among these proteins, we further characterized a lipoxygenase homology gene CaLOP (Capsicum annuum lipoxygenase homology protein), which has a 730 bp cDNA sequence and encodes 184 amino acids. CaLOP contains a lipoxygenase homology (PLAT/LH2) domain. A database search revealed that CaLOP shares significant sequence identity with putative orthologs in other plants. CaLOP was constitutively expressed in root tissue, and was upregulated in response to TMV-P0, TMV-P2, and PepMoV viruses, as well as Phytophthora capsici. To examine the biological function of this gene in plant development, we performed in planta knockdown assays using the Tobacco rattle virus-based gene-silencing system. Silencing CaLOP in pepper resulted in abnormal growth and severe developmental disorders, including lethal phenotypes, compared with controls. Consequently, our results suggest that CaLOP may be important in plant growth and development, and pathogen defense.

Keywords

Capsicum

lipoxygenase-homology protein

secreted protein

VIGS

식물 세포벽 및 세포 사이에 존재하는 분비단백질은 식물 생육과 발달뿐만 아니라 다양한 환경 스트레스 조절 기작에 관여한 다. 선행 연구에서 고추와 병원균의 상호작용을 통해 분비된 고추 유래 분비단백질의 대량 동정을 통해 방어 기제 연구가 진행 되었다. 이 중 병원균 상호작용 분비 단백질의 작물 발달 및 생육 관련 연구를 위해 CaLOP 유전자를 선발하였다. CaLOP 유전 자는 PLAT/LH2 도메인을 가지고 있었으며, 같은 가지과 식물인 토마토, 감자, 담배 이외에도 애기장대와 벼에서 이종 상동성 유전자들을 확인할 수 있었다. 발현 양상을 살펴보면, CaLOP 유전자는 뿌리 조직에서 강하게 발현되었으며, 고추 역병뿐만 아 니라 TMV-P0, TMV-P1,2 및 PepMoV 바이러스와의 상호작용 과정의 특정 시기에 따라 발현이 증가하였다. 식물 생육 및 발 달에서의 기능을 확인하기 위해, 유전자의 침묵 현상을 유도하여 CaLOP 유전자의 발현을 억제한 결과 대조군에 비해 엽면적 이 작아지고, 초장이 감소하는 등의 비정상적 생육과 생장점의 고사 등의 심각한 발달장애가 나타났다. 이를 통해 CaLOP 유전 자는 병원균에 대항하는 방어기제 이외에도 식물의 생육과 발달에 중요한 기능을 하는 유전자라는 것을 확인할 수 있었다.

키워드

고추

리포옥시게네이즈 유사 유전자

분비단백질

유전자 침묵 현상

References

Hugot K, Riviere MP, Moreilhon C, Dayem MA, Cozzitorto J, Arbiol G, Barbry P, Weiss C, Galiana E (2004) Coordinated regulation of genes for secretion in tobacco at late developmental stages: association with resistance against oomycetes. Plant Physiol 134:858-870. doi:10.1104/pp.103.034173

Kang WH, Kim S, Lee HA, Choi D, Yeom SI (2016) Genome-wide analysis of Dof transcription factors reveals functional characteristics during development and response to biotic stresses in pepper. Sci Rep 6:33332. doi:10.1038/srep33332

Kim HJ, Lee HR, Han JH, Yeom SI, Harn CH, Kim BD (2008) Marker production by PCR amplification with primer pairs from conserved sequences of WRKY genes in chili pepper. Mol Cells 25:196-204. doi:10.1007/s10059-009-0002-6

Kim S, Park M, Yeom SI, Kim YM, Lee JM, Lee HA, Seo E, Choi J, Cheong K, et al (2014) Genome sequence of the hot pepper provides insights into the evolution of pungency in Capsicum species. Nat Genet 46:270-278. doi:10.1038/ng.2877

Lee HA, Yeom SI (2015) Plant NB-LRR proteins: tightly regulated sensors in a complex manner. Brief Funct Genomics 14:233-242. doi:10.1093/bfgp/elv012

Lee HR, Kim KT, Kim HJ, Han JH, Kim JH, Yeom SI, Kim HJ, Kang WH, Shi J, et al (2011) QTL analysis of fruit length using rRAMP, WRKY, and AFLP markers in chili pepper. Hortic Environ Biotechnol 52:602-613. doi:DOI 10.1007/s13580-011-0002-2

Lee SJ, Saravanan RS, Damasceno CM, Yamane H, Kim BD, Rose JK (2004) Digging deeper into the plant cell wall proteome. Plant Physiol Biochem 42:979-988. doi:10.1016/j.plaphy.2004.10.014

Schmelzer E (2002) Cell polarization, a crucial process in fungal defence. Trends Plant Sci 7:411-415. doi:10.1016/S1360-1385 (02)02307-5

Schulze-Lefert P (2004) Knocking on heaven's wall: pathogenesis of and resistance to biotrophic fungi at the cell wall. Curr Opin Plant Biol 7:377-383. doi:10.1016/j.pbi.2004.05.004

Seo E, Kim S, Yeom SI, Choi D (2016) Genome-wide comparative analyses reveal the dynamic evolution of nucleotide-binding leucine-rich repeat gene family among solanaceae plants. Front Plant Sci 7:1205. doi:10.3389/fpls.2016.01205

Seo E, Yeom SI, Jo S, Jeong H, Kang BC, Choi D (2012) Ectopic expression of Capsicum-specific cell wall protein Capsicum annuum senescence-delaying 1 (CaSD1) delays senescence and induces trichome formation in Nicotiana benthamiana. Mol Cells 33:415-422. doi:10.1007/s10059-012-0017-2

Shin R, An JM, Park CJ, Kim YJ, Joo S, Kim WT, Paek KH (2004) Capsicum annuum tobacco mosaic virus-induced clone 1 expression perturbation alters the plant's response to ethylene and interferes with the redox homeostasis. Plant Physiol 135:561-573. doi:10.1104/pp.103.035436

Shin R, Kim MJ, Paek KH (2003) The CaTin1 (Capsicum annuum TMV-induced clone 1) and CaTin1-2 genes are linked head-to-head and share a bidirectional promoter. Plant Cell Physiol 44:549-554. doi:10.1093/pcp/pcg069

Tomita R, Sekine K-T, Mizumoto H, Sakamoto M, Murai J, Kiba A, Hikichi Y, Suzuki K, Kobayashi K (2011) Genetic basis for the hierarchical interaction between tobamovirus spp. and l resistance gene alleles from different pepper species. Mol Plant Microbe Interact 24:108-117. doi:10.1094/MPMI-06-10-0127

Yamane H, Lee SJ, Kim BD, Tao R, Rose JK (2005) A coupled yeast signal sequence trap and transient plant expression strategy to identify genes encoding secreted proteins from peach pistils. J Exp Bot 56:2229-2238. doi:10.1093/jxb/eri222

Yang H-B, Liu W-y, Kang W-H, Kim J-H, Cho HJ, Yoo J-H, Kang B-C (2012) Development and validation of L allele-specific markers in Capsicum. Mol Breed 30:819-829. doi:10.1007/s11032-011-9666-7

Yeom SI, Baek HK, Oh SK, Kang WH, Lee SJ, Lee JM, Seo E, Rose JK, Kim BD, et al (2011) Use of a secretion trap screen in pepper following phytophthora capsici infection reveals novel functions of secreted plant proteins in modulating cell death. Mol Plant Microbe Interact 24:671-684. doi:10.1094/MPMI-08-10-0183

Yeom SI, Seo E, Oh SK, Kim KW, Choi D (2012) A common plant cell-wall protein HyPRP1 has dual roles as a positive regulator of cell death and a negative regulator of basal defense against pathogens. Plant J 69:755-768. doi:10.1111/j.1365-313X.2011.04828.x

Information

Publisher :KOREAN SOCIETY FOR HORTICULTURAL SCIENCE
Publisher(Ko) :한국원예학회
Journal Title :Horticultural Science and Technology
Journal Title(Ko) :원예과학기술지
Volume : 36
No :2
Pages :292-302
Received Date : 2017-04-04
Revised Date : 2017-07-05
Accepted Date : 2017-06-27
DOI :https://doi.org/10.12972/kjhst.20180029

Horticultural Science and Technology 원예과학기술지 ISSN:1226-8763(Print) 2465-8588(Online)

All Issue