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. 2010 Feb;231(3):767-77.
doi: 10.1007/s00425-009-1087-z. Epub 2009 Dec 24.

The MADS-domain protein MPF1 of Physalis floridana controls plant architecture, seed development and flowering time

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The MADS-domain protein MPF1 of Physalis floridana controls plant architecture, seed development and flowering time

Chaoying He et al. Planta. 2010 Feb.

Abstract

Floral and vegetative development of plants is dependent on the combinatorial action of MADS-domain transcription factors. Members of the STMADS11 subclade, such as MPF1 of Physalis, are abundantly expressed in leaves as well as in floral organs, but their function is not yet clear. Our studies with transgenic Arabidopsis that over-express MPF1 suggest that MPF1 interacts with SOC1 to determine flowering time. However, MPF1 RNAi-mediated knockdown Physalis plants revealed a complex phenotype with changes in flowering time, plant architecture and seed size. Flowering of these plants was delayed by about 20% as compared to wild type. Expression of PFLFY is upregulated in the MPF1 RNAi lines, while PFFT and MPF3 genes are strongly repressed. MPF1 interacts with a subset of MADS-domain factors, namely with PFSOC1 in planta, and with PFSEP3 and PFFUL in yeast, supporting a regulatory role for this protein in flowering. The average size of seeds produced by the transgenic MPF1 RNAi plants is increased almost twofold. The height of these plants is also increased about twofold, but most axillary buds are stunted when compared to controls. Taken together, this suggests that members of the STMADS11 subclade act as positive regulators of flowering but have diverse functions in plant growth.

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Figures

Fig. 1
Fig. 1
Analysis of over-expressing MPF1 transgenic Arabidopsis plants. a, b Phenotype of a typical 35S:MPF1 transgenic line in comparison to wild-type Arabidopsis grown under long-day (LD) and short-day (SD) conditions, respectively. c RT-PCR analysis of MPF1 gene expression in transgenic Arabidopsis. The upper graph shows data from real-time RT-PCR. The PP2A gene was used as a loading control and for normalization. Error bars represent ±SD (n = 3). The lower panel shows semi-quantitative RT-PCR analysis. d Non-lethal β-galactosidase assays of possible interactions of STMADS11, MSM1 and MPF1 with several other MADS-box proteins (listed on top). The empty vector pGBKT7 and AP2 from Arabidopsis were used as controls
Fig. 2
Fig. 2
MPF1-like gene expression. a Analysis of STMADS11 and MPF1 expression by Northern blotting. ACTIN was used as a loading control. Total RNAs were extracted from leaves (L), sepals, petals, stamens and carpels from buds (B), small buds (sB), large buds (lB), flowers (F), young flowers (yF) or mature flowers (mF), and young fruits (yFr), large fruits (lFr) and old fruits (oFr), respectively. b Functional promoter analysis: GUS expression pattern in STMADS11:GUS (upper) and MPF1:GUS (lower) transgenic Arabidopsis lines. GUS signals are in leaf, inflorescence (flower buds), mature flower and silique, but not in ovary and seeds (from left to right)
Fig. 3
Fig. 3
Characterization of putative target genes and interacting proteins of MPF1 in Physalis. a Non-lethal β-galactosidase assay for interaction between MPF1, the full-length PFSOC (1–219) and its truncated derivatives (gene and sizes are indicated) in yeast. b, c Bimolecular fluorescence complementation (BiFC) assay after agroinfiltration of MPF1 with PFSOC1 into leaf epidermal cells of P. floridana. Fluorescence (green signal) reveals interaction between MPF1 with PFSOC1 in planta. The relevant combinations of complementation constructs are indicated in the panels. Arrows indicate the nuclei
Fig. 4
Fig. 4
Expression of MPF1 and flowering time of 35S:MPF1-RNAi transgenic Physalis lines. a Expression of MPF1 in 35S:MPF1-RNAi lines. MPF1 expression was measured in the 12 knockdown (KO) transgenic lines using real-time RT-PCR, and expression levels were normalized relative to wild-type (WT). Error bars represent ±SD (n = 3). b Flowering time of the transgenic 35S:MPF1-RNAi lines and wild-type Physalis. Flowering time is given in days to anthesis. The black column represents wild type and the gray columns represent individual transgenic lines. Days 0–30 were condensed to allow magnification of days 30–70 for apparent differences. Error bars represent ±SD (n = 4). c The expression levels of the putative flowering-time genes (from left to right) in wild-type Physalis (black column) and the transgenic knockdown lines KO2 (dark gray), KO3 (middle gray), KO4 (light gray) and KO10 (white). Plant lines were grown for 28 days under long-day conditions. The expression level of each gene was normalized relative to wild type (black column). Error bars represent ±SD (n = 3)
Fig. 5
Fig. 5
Plant architecture, seed size and seed weight in transgenic Physalis. a Plant architecture. The most severely affected 35S:MPF1-RNAi lines (KO2, KO3, KO5, KO6, KO9 and KO10) were selected, and their vegetative development was examined when the first flower opened. Average lengths of the elements indicated were determined for each of the six lines and normalized relative to wild type. The data points shown as black dots indicate relative overall lengths. Internode number is given by gray squares and internode length by gray triangles. MS is the main stem, CS1 and CS2 indicate cotyledonous axillary side shoots and LS1 to LS7 are axillary side shoots of the main stem. b Schematic representation of growth habit of 35S:MPF1-RNAi and wild-type plants. DB is first dichotomous branch (other abbreviations as in a); the black dot indicates the first flower. Arrowheads show the direction of growth. c Comparison of seed size in wild type and the KO2 line. d Seed weight in transgenic Physalis lines (Fig. 4a)

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