HMG蛋白是核蛋白第二多的蛋白家族，在調控DNA彎曲和染色體構型扮演重要角色。在阿拉伯芥中， 16個HMG蛋白被分為4類：HMGB, SSRP, 3xHMG-Box, 和ARID-HMG，而HMGB15是屬於ARID-HMG類別。在先前的研究中，阿拉伯芥中HMGB15表現量降低時會造成花粉管延遲生長，並且會導致果莢短小及子代數目減少。在本研究中我們發現HMGB15會和組蛋白去乙醯化酶HDA6在細胞核中有交互作用。藉由酵母菌雙雜合系統實驗進一步證明 HMGB15蛋白會和HDA6蛋白的C端有交互作用。同時，hmgb15的突變株在短日照下有早開花的表型，而HMGB15過表達殖株會有晚開花的表型。通過檢測hmgb15突變株中及過表達植物中基因表達，我們發現相對於野生型，參與開花調控的下游基因表達在hmgb15突變株和過表達植物也都有受到影響。另外，當在hda6突變株中過量表達HMGB15時會造成早開花的表型，說明HMGB15作用在HDA6下游。綜合上述結果，我們可以知道HMGB15會和HDA6有交互作用並且調控植物開花時間。

HMG-box containing proteins are the second most abundant nuclear protein family in plants cells. They play an important role in modulation of DNA bending and chromosome architecture. In Arabidopsis, 16 HMG proteins can be classified into 4 classes: HMGB, SSRP, 3xHMG-Box, and ARID-HMG. AtHMB15 belongs to the ARID-HMG class. Previous studies showed that knocking down AtHMGB15 expression via a Ds insertion caused retarded pollen tube growth, leading to a significant reduction in the seed set. HDA6 is a RPD3-type histone deacetylase involved in flowering and hda6 mutants display a late flowering phenotype. In this study, it was showed that HDA6 interacts with HMGB15 in the nucleus. Moreover, HMGB15 interacts with the C-terminus of HDA6 in yeast two hybrid assays. Compared to wild type, hmgb15 mutants flowered early under short day conditions while HMGB15 overexpressing plants showed late flowering. The expression levels of the genes involved in flowering time control were changed in hmgb15 and HMGB15 overexpressing plants compared with wild type. Furthermore, overexpressing HMGB15 in the hda6 mutant resulted in an early flowering phenotype, indicating that HMGB15 may act downstream of HDA6 in flowering time control. Taken together, our results indicate that HMGB15 interacts with HDA6 and is involved in flowering time control in Arabidopsis.

致謝.... I
中文摘要........ II
Abstract....... III
List of Tables......... VI
List of Figures........ VII
List of Supplementary Figures.......... IX
List of abbreviations.......... X
Introduction... 1
High Mobility Group (HMG) proteins..... 1
Structural variability and function of plant HMGB proteins....... 2
Histone deacetylases in Arabidopsis.... 4
Materials and Methods.. 5
Plant materials........ 6
Quick DNA extraction... 6
RNA isolation.......... 7
DNase treatment........ 9
Quantitative RT-PCR analysis........... 9
Chromatin immunoprecipitation assays... 11
Bimolecular Fluorescence Complementation (BiFC) assays ........................................20
Transfection of tobacco leaves by Agrobacterium (For BiFC assays)................................ 23
Yeast two-hybrid assays................ 24
Western blot assays:................... 26
Results................................ 28
Phylogenic analysis and sequence comparison of HMG proteins............................... 28
The expression pattern of ARID-HMG genes....... 28
Subcellular localization of HMGB15 in Arabidopsis ........................................29
HMGB15 interacts with HDA6............. 29
Identification of interaction domains of HDA6 and HMGB15 ........................................30
Identification of homozygous T-DNA insertion mutants of HMGB15................................. 31
hmgb15 mutant plants display early flowering phenotypes ........................................31
HMGB15 is involved in flowering time control in Arabidopsis............................ 32
HMGB15 overexpressing plants show delayed flowering ........................................32
HDA6 and FLD regulate the expression of HMGB15................................. 33
Early flowering phenotype of hda6-6/HMGB15-OE3.....................................34
Discussion............................. 35
HMGB15 interacts with the histone deacetylase HDA6 ........................................35
HMGB15 regulates flowering in Arabidopsis............................ 37
Figures.................................39
Supplementary Figure................... 63
References............................. 70

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