林詩舜 助理教授

E-mail︰linss01@ntu.edu.tw

專長︰植物分子生物學 / 植物分子病毒學 / 植物生物技術

連絡：生物科技所 411

Tel︰886-2-3366-6023

Fax︰886-2-3366-6001

個人網站：http://homepage.ntu.edu.tw/~linss01

2001  博士  國立中興大學農業生物技術研究所

1995  學士  國立中興大學園藝學

助理教授 2008~now  國立臺灣大學

博士後研究 2003~2008  美國紐約洛克菲勒大學

博士後研究 2003  國立中興大學

2004~2006  教育部博士後研究獎學金

2001 中華民國翡陶斐榮譽學會

2001~1997  國科會博士班學生獎學金

1997~1996  國科會碩士班學生獎學金

1. Special topic of plant biotechnology
   
   
2. Advance small RNA and mechanism of gene silencing
   
   
3. Epigenetics
   
   
4. Frontiers in biotechnology
   
   
5. Plant molecular virology
   

Our Laboratory is major studying (1) the application of artificial miRNA (amiRNA); (2) the function of viral suppressor; and (3) plant cross-protection using genetic, molecular, cellular, biochemical, genomic, and proteomic approaches.

Artificial miRNA (amiRNA)
AmiRNA technology can be applied on resistance of plant virus and precisely cleavage plant endogenous mRNA (Fig.1). This approach can increase the bio-safety of genome modified organ (GMO) products, which are created by RNA-mediated resistance or RNA interfering (RNAi) approaches using viral gene segments. The designing of the amiRNA is according to the endogenous precursor of the miRNA as a backbone and modified the miRNA/miRNA* sequence to the amiRNA/amiRNA* sequence by PCR mutagenesis. The modified of the amiRNA precursor will be constructed into a binary vector and then transferred into plant for the amiRNA over-expression (Fig. 1).

There are several advantages to the use of amiRNA strategy. (1) Only one stable small RNA (21-nt) is produced by the amiRNA approach, and its sequence can be chosen to reduce off-target effects. (2) The amiRNA strategy also minimizes potential risks for bio-safety concern and also reduces any possible negative environmental impact. (3) Broad-spectrum resistance to several viruses can also be achieved by co-expression of appropriately designed multiple amiRNAs.

Figure 1. Design of amiRNA precursors and amiRNA-mediated of viral resistance. (A) A structure of pre-miR159a presented as a hairpin. Primers containing the amiRNA or amiRNA* sequences were used to replace miR159 and miR159* sequence in pre-mRNA. The pre-amiRNA159 was subcloned into pENTR vector and then moved downstream of a 35S promoter in a binary vector and transferred into plant. (B) Transgenic Arabidopsis expressing amiR-P69159/amiR-HC-Pro159 are resistant to Turnip yellow mosaic virus (TYMV) and Turnip mosaic virus (TuMV) infection. (Nature Biotechnology 24:1420-1428)

Viral gene silencing suppressor
We are also focusing on the function of viral gene silencing suppressors and mechanism of post-transcription gene silencing (PTGS). In Arabidopsis, there are 4 Dicer-like proteins (DCLs) and 10 Agronaues (AGO). These DCLs and AGOs are cross-interacted each other and created various gene silencing pathways. Various DCL-AGO recombinations have its own specific function, ex. DCL1-AGO1 complexes are involved in miRNA pathway; DCL4 interacted with unknown function of AGO and response for virus-induce gene silencing (VIGS) pathway. We found several important amino acids that are necessary and sufficient to suppress various gene silencing pathways and these suppressor mutants will help to understand their mechanisms. In addition, we are studying on protein-protein interaction between viral suppressor and endogenous gene of host. We would like to identify uncover novel genes which regulate and control gene silencing pathways (Fig. 2).


Figure 2. The Arabidopsis wild-type (leaf) or expressing viral suppressor (P1/HC-Pro) gene (right).

Plant cross-protection
Cross-protection has been used for the control of virus disease. This resistance is induced on a host plant by infection with an attenuated strain of a virus (termed as protective virus) (Fig. 3). As a result of this infection, the infected plant is rendered to subsequent infections by a closely related strain of virus (termed as challenge virus). The key point for cross-protection is that that the protective virus, also named the attenuated virus or mild strain, induced mild or no symptoms at all on the host. Once the mild strain has been inoculated it can trigger strain- or sequence-specific resistance against the challenge virus. Many researches have found that mutations on viral silencing suppressors can attenuate severe symptoms, e.g. mutations in the silencing suppressor HC-Pro from potyvirus.


Figure 3. The strategies of plant virus resistance.

The phenomenon of cross-protection is very similar to the immune response in animals, which is induced by attenuated viral vaccines. However, plants do not possess a similar immune system and the mechanism of cross-protection remains unclear. It is possible that cross-protection is mediated by protein or by RNA or a combination of mechanism (Fig. 4).


Figure 4. A model to explain the cross protection in squash plants provide by the constructed Zucchini yellow mosaic virus (ZYMV)-GAC and ZYMV-GAB mutants and the naturally collected mild strain ZYMV WK against the severe strain ZYMV. (Phytopathology 97:287-296)

We believe future investigations on the mechanism of cross-protection will help to uncover novel mechanisms in virus-plant interaction and provide knowledge for the development of more efficient strategies with improved bio-safety for virus resistance in the field.

1. Wu, H.W.*, Lin, S.S.*, Chen, K.C., Yeh, S.D., Chua, N.H. 2010. Discriminating mutations of HC-Pro of Zucchini yellow mosaic virus with differential effects on small RNA pathways involved in viral pathogenicity and symptom development. MPMI. 23:17-28 (*co-first author)
2. 
   Lin, S.S.*, Wu, H.W.*, Elena, S.F., Chen, K.C., Niu, Q.W., Yeh, S.D., Chen, C.C., Chua, N.H. 2009. Molecular evolution of a viral non-coding sequence under the selective pressure of amiRNA-mediated silencing. PLoS Pathog. 5(2):e1000312. Doi:10.1371/journal.ppat.1000312 (*co-first author)
3. 
   Lin, S.S., Martin, R., Mongrand, S., Vandenabeels, S., Chen, K.C., Jang, I.C., Chua, N.H.  2008. RING1 E3 ligase localizes to plasma membrane lipid raft to trigger FB1-induced programmed cell death in Arabidopsis. Plant J. 56:550-561
   
   
4. Lin, S.S., Henriques, R., Wu, H.W., Niu, Q.W., Yeh, S.D., and Chua, N.H.  2007. Strategies and mechanisms of plant virus resistance.  Plant Biotechnol. Rep. 1:125-134 (Review)
   
   
5. Lin, S.S., H.W. Wu, F.-J. Jan, Hou, R.F., and Yeh, S.D. 2007. Modifications of the helper component-protease of Zucchini yellow mosaic for generation of attenuated mutants for cross protection against severe infection. Phytopathology 97:287-296 (Cover photo)
   
   
6. Niu, Q.W.*, Lin, S.S.*, Reyes J.L., Chen, K. C., Wu, H.W., Yeh, Y.D., and Chua, N.H. 2006. Expression of artificial microRNAs in transgenic Arabidopsis thaliana confers virus resistance Nat. Biotech. 24:1420-1428 (*co-first author)
   
   
7. Chiang, C.H., Lee, C.Y., Wang, C.H., Jan, F.J., Lin, S.S., Chen, T.C., Raja, J.A.J, and Yeh, S.D.  2007.  Genetic analysis of an attenuated Papaya ringspot virus strain applied for cross-protection.  Eur. J. Plant Pathol. 118:333-348
   
   
8. Chen, C.C., Chen, T.C., Raja, J.A., Chang, C.A., Chen, L.W., Lin, S.S., Yeh, S.D. 2007. Effectiveness and stability of heterologous proteins expressed in plants by Turnip mosaic virus vector at five different insertion sites. Virus Res. 130:210-227
   
   
9. Zhang, X., Yuan, Y.U., Pei, Y., Lin, S.S., Tuschl, T., Patel, D.J., and Chua, N.H. 2006. Cucumber mosaic virus-encoded 2b suppressor inhibits Arabidopsis AGO1 cleavage activity to counter plant defense. Gene Dev. 20:3255-3268
   
   
10. Zhang, X., Henriques, R., Lin, S.S., Niu, Q.W., Chua, N.H. 2006. Agrobacterium-mediated transformation of Arabidopsis thaliana using the floral-dip method.  Nat. Protocol. 1:641-646.
    
    
11. Hsu, C.H., Lin, S.S., Liu, F.L., Su,W.C., and Yeh, S.D. 2004. Oral administration of mite allergen expressed by zucchini yellow mosaic virus in cucurbit species downregulates allergen-induced airway inflammation and IgE synthesis. J. Allergy Clin Immunol. 113:1079-1085.
    
    
12. Lin, S. S., Hou, R. F., and Yeh, S. D. 2002. Construction of in vitro and in vivo infectious transcripts of a Taiwan strain of Zucchini yellow mosaic virus. Bot. Bull. Acad. Sin. 43:261-269.
    
    
13. Lee, K.C., Lin, S.S., Yeh, S.D., and Wong, S.M. 2002. Interations between nuclear inclusion protein a (NIa) and Nuclear Inclusion Protein b (NIb) of Zucchini yellow mosaic virus and Papaya ringspot virus. Plant Prot. Bull. 11:79-86.
    
    
14. Lin, S. S., Hou, R. F., and Yeh, S. D. 2001. Complete genome sequence and genetic organization of a Taiwan isolate of Zucchini yellow mosaic virus. Bot. Bull. Acad. Sin. 42: 243-250.
    
    
15. Chu F. H., Chao, C. H., Peng, Y. C. Lin, S. S., Chen, C. C., Yeh, S. D. 2001. Serological and molecular characterization of Peanut chlorotic fan-spot virus, a new species of the genus Tospovirus. Phytopathology 91: 856-863.
    
    
16. Lin, S.S., Hou, R.F., and Yeh, S.D. 2000. Heteroduplex mobility and sequence analyses for assessment of variability of Zucchini yellow mosaic virus. Phytopathology 90: 228-235.
    
    
17. Huang, C.H., Lin, S.S., and Yeh, S.D. 2000. Sequence analysis of the coat protein gene of Zucchini yellow mosaic virus isolated from diseased woody fruit of bitter gourd. Plant Pathol. Bull. 10:11-18.
    
    
18. Lin, S.S., Hou, R.F., Huang, C.H., and Yeh, S.D. 1998. Characterization of Zucchini yellow mosaic virus (ZYMV) isolates collected from Taiwan by host reactions, serology, and RT-PCR. Plant Prot. Bull. 40: 163-176.

CONGRESS PUBLICATION

1. Lin, S.S. Wu, H.W., Chen, K.C., Yeh, S.D., and Chua, N.H. 2009. Application of artificial microRNA. 10th AEARU workshop on molecular biology and biotechnology. (Taipei, Nov. 11-13)
   
   
2. Wu, H.W., Lin, S.S., Chen, K.C., Yeh, S.D., Chua, N.H. 2008. Discriminating mutations of HC-Pro with differential effects on small RNA pathways. Abstracts of XIV. International Congress of Virology for IUMS 2008, (Istanbul, Aug, 10-15)
   
   
3. Lin, S.S., Vandenabeele, S., and Chua, N.H. 2004. Regulation of programmed cell death by an Arabidopsis RING motif protein. Abstracts of Disease resistance and related signaling mechanisms in plants on Centre for International Meetings on Biology, Spain
   
   
4. Yeh, S.D., Hou, R.F., and Lin, S.S. 2002. Construction of mild strains of Zucchini yellow virus by point mutations in the helper component-protease gene and evaluation of their cross-protection effectiveness. The World of Microbes, p.15. Abstracts of XIIth International Congress of Virology (Paris, July 27-Aug 1).
   
   
5. Chen, T.C., Huang, C.W., Lin Y.H., Liu, F.L., Lin. S.S., and Yeh, S.D. 2002. Efficient expression of the NSs and N proteins of thrips-borne Watermelon silver mottle tospovirus in aquash by Zucchini yellow mosaic potyvirus vector. The World of Microbes, p. 378. Abstracts of XIIth International Congress of Virology (Paris, July 27-Aug 1)
   
   
6. Lin, S.S., Hsu, C.H., Huang, X.W., Hou, R.F., Yeh, S.D. 2001. Development of Zucchini yellow mosaic virus as highly efficient, stable vector for expressing foreign proteins in cucurbit species.  Abstracts of Second International Symposium on Biotechnology of Tropical and Subttropical Species. p.72 (Academica Sinica, November 5-9, 2001)
   
   
7. Lin, S.S., Hsu, C.H., Huang, X.W., Hou, R.F., and Yeh, S.D. 2001 Development of a Taiwan strain of Zucchini yellow mosaic virus as a highly efficient, stable vector for expressing foreign proteins in cucurbit species. 3th Cross-Strait Symposium on Plant Molecular Biology and Biotechnology. p.21 (Hong Kong, August 5-11)
   
   
8. Huang, C.W., Chen, T.C., Lin, Y.S., Liu, F.L., Lin, S.S., and Yeh, S.D. 2001. Expression of the N and NSs genes of Watermelon silver mottle virus in a squash by the Zucchini yellow mosaic virus vector. Plant Path. Bull. 10:216 (Pingtung, 2001, Dec 8-9).
   
   
9. Lin, S.S., Hou, R.F., and Yeh, S.D. Construction of a viral vector from the in vivo infectious clone of Zucchini yellow mosaic virus for expressing foreign proteins in plants. 6th International Congress of Plant Molecular Biology, Quebec, Canada, Book of Abstracts S30-48. June 18-24, 2000
   
   
10. Lin, S.S., Hou, R.F., and Yeh, S.D. Heteroduplex mobility analysis and coat protein variability of Zucchini yellow mosaic virus isolates from Taiwan. 1998 Joint annual meeting American Phytopathological Society and Entomological Society of America. LasVegas, NV, November 8-12, 1998