Transcriptome Analysis of Triple Mutant for OsMADS62, OsMADS63, and OsMADS68 Reveals the Downstream Regulatory Mechanism for Pollen Germination in Rice (Oryza sativa)

The MADS (MCM1-AGAMOUS-DEFFICIENS-SRF) gene family has a preserved domain called MADS-box that regulates downstream gene expression as a transcriptional factor. Reports have revealed three MADS genes in rice, OsMADS62, OsMADS63, and OsMADS68, which exhibits preferential expression in mature rice pollen grains. To better understand the transcriptional regulation of pollen germination and tube growth in rice, we generated the loss-of-function homozygous mutant of these three OsMADS genes using the CRISPR-Cas9 (clustered regularly interspaced short palindromic repeats-CRISPR associated protein 9) system in wild-type backgrounds. Results showed that the triple knockout (KO) mutant showed a complete sterile phenotype without pollen germination. Next, to determine downstream candidate genes that are transcriptionally regulated by the three OsMADS genes during pollen development, we proceeded with RNA-seq analysis by sampling the mature anther of the mutant and wild-type. Two hundred and seventy-four upregulated and 658 downregulated genes with preferential expressions in the anthers were selected. Furthermore, downregulated genes possessed cell wall modification, clathrin coat assembly, and cellular cell wall organization features. We also selected downregulated genes predicted to be directly regulated by three OsMADS genes through the analyses for promoter sequences. Thus, this study provides a molecular background for understanding pollen germination and tube growth mediated by OsMADS62, OsMADS63, and OsMADS68 with mature pollen preferred expression.

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BcSKS11, a SKS gene from pollinated pistils of Chinese cabbage pak choi, is expressed continuously during the male gametophyte development, pollen germination and pollen-tube growth processes

Australian Journal of Botany ◽

10.1071/bt12218 ◽

2013 ◽

Vol 61 (6) ◽

pp. 446

Author(s):

Zhixian Zhang ◽

Jiashu Cao ◽

Lin Qiu ◽

Jingjing Jiang ◽

Baohua Wu

Keyword(s):

Cell Wall ◽

Pollen Tube ◽

Chinese Cabbage ◽

Pollen Tube Growth ◽

Pollen Germination ◽

In Situ Hybridisation ◽

Pollen Grains ◽

Tube Growth ◽

Mature Pollen ◽

Pak Choi

Cell wall-related genes and proteins are crucial for fertilisation-related events such as pollen germination and pollen-tube growth. Recent studies have focussed on the molecular mechanism of cell-wall synthesis and regulation of these processes; however, limited information is available on the mechanism and modification of the crucial cell-wall materials. We isolated a gene that is potentially involved in cell-wall metabolism from pollinated pistils of the male sterile line ‘Bcajh97-A’ in Chinese cabbage pak choi (Brassica campestris SKewed5 Similar 11, BcSKS11). BcSKS11 has several features of SKewed5 Similar (SKS) proteins, which are structures that are similar to plant multicopper oxidase but lack the 10 necessary residues of two copper-binding sites. A phylogenetic tree constructed from 49 SKS proteins found four clades, with BcSKS11 being in a clade with tobacco sequences. Reverse transcription–polymerase chain reaction and in situ hybridisation revealed that BcSKS11 was specifically expressed in uninucleate microspores, mature pollen grains and growing pollen tubes of pollinated pistils. These results suggest that BcSKS11 might be stored in mature pollen grains, in preparation for pollen germination and pollen-tube growth.

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Transcriptome and metabolome profiling provide insights into molecular mechanism of pseudostem elongation in banana

BMC Plant Biology ◽

10.1186/s12870-021-02899-6 ◽

2021 ◽

Vol 21 (1) ◽

Author(s):

Guiming Deng ◽

Fangcheng Bi ◽

Jing Liu ◽

Weidi He ◽

Chunyu Li ◽

...

Keyword(s):

Cell Wall ◽

Cell Elongation ◽

Molecular Mechanisms ◽

Specific Gene ◽

Wild Type ◽

Banana Plant ◽

Cell Wall Modification ◽

Dwarf Cultivar ◽

Important Trait ◽

Metabolome Profiling

AbstractBackgroundBanana plant height is an important trait for horticultural practices and semi-dwarf cultivars show better resistance to damages by wind and rain. However, the molecular mechanisms controlling the pseudostem height remain poorly understood. Herein, we studied the molecular changes in the pseudostem of a semi-dwarf banana mutant Aifen No. 1 (Musaspp. Pisang Awak sub-group ABB) as compared to its wild-type dwarf cultivar using a combined transcriptome and metabolome approach.ResultsA total of 127 differentially expressed genes and 48 differentially accumulated metabolites were detected between the mutant and its wild type. Metabolites belonging to amino acid and its derivatives, flavonoids, lignans, coumarins, organic acids, and phenolic acids were up-regulated in the mutant. The transcriptome analysis showed the differential regulation of genes related to the gibberellin pathway, auxin transport, cell elongation, and cell wall modification. Based on the regulation of gibberellin and associated pathway-related genes, we discussed the involvement of gibberellins in pseudostem elongation in the mutant banana. Genes and metabolites associated with cell wall were explored and their involvement in cell extension is discussed.ConclusionsThe results suggest that gibberellins and associated pathways are possibly developing the observed semi-dwarf pseudostem phenotype together with cell elongation and cell wall modification. The findings increase the understanding of the mechanisms underlying banana stem height and provide new clues for further dissection of specific gene functions.

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Responsiveness to exogenous cAMP of a Saccharomyces cerevisiae strain conferred by naturally occurring alleles of PDE1 and PDE2.

Genetics ◽

10.1093/genetics/135.2.321 ◽

1993 ◽

Vol 135 (2) ◽

pp. 321-326 ◽

Cited By ~ 2

Author(s):

H Mitsuzawa

Keyword(s):

Saccharomyces Cerevisiae ◽

Slow Growth ◽

Loss Of Function ◽

Wild Type ◽

Triple Mutant ◽

Apparent Absence ◽

Camp Pathway ◽

Naturally Occurring ◽

Elevated Activity ◽

Growth Phenotype

Abstract The Saccharomyces cerevisiae strain P-28-24C, from which cAMP requiring mutants derived, responded to exogenously added cAMP. Upon the addition of cAMP, this strain showed phenotypes shared by mutants with elevated activity of the cAMP pathway. Genetic analysis involving serial crosses of this strain to a strain with another genetic background revealed that the responsiveness to cAMP results from naturally occurring loss-of-function alleles of PDE1 and PDE2, which encode low and high affinity cAMP phosphodiesterases, respectively. In addition, P-28-24C was found to carry a mutation conferring slow growth that lies in CYR1, which encodes adenylate cyclase, and the slow growth phenotype caused by the cyr1 mutation was suppressed by the pde2 mutation. Therefore P-28-24C is fortuitously a pde1 pde2 cyr1 triple mutant. Responsiveness to cAMP conferred by pde mutations suggests that S. cerevisiae cells are permeable to cAMP to some extent and that the apparent absence of effect of exogenously added cAMP on wild-type cells is due to immediate degradation by cAMP phosphodiesterases.

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Impact of Manganese on Pollen Germination and Tube Growth in Lily

Acta Agrobotanica ◽

10.5586/aa.746 ◽

2021 ◽

Vol 74 ◽

Author(s):

Thomas Sawidis ◽

Gülriz Baycu ◽

Elżbieta Weryszko-Chmielewska ◽

Aneta Sulborska

Keyword(s):

Pollen Tube ◽

Pollen Germination ◽

Surrounding Medium ◽

Pollen Grains ◽

Lilium Longiflorum ◽

Tube Growth ◽

Low Concentrations ◽

Main Effect

Abstract In vitro culture of Lilium longiflorum pollen grains was carried out to determine the role of manganese in pollen germination and pollen tube growth. Pollen germination was adversely affected by the presence of manganese (>10 −8 M), whereas low concentrations (10 −12 –10 −10 M) stimulated the process. Manganese caused morphological anomalies during tube growth, characterized by irregular pollen tube thickening and swollen tips. The main effect was the anomalous cell wall formation at the tip, in which the presence of several organelles reduced the number of secretory vesicles. A loose network of fibrillar material and spherical aggregates, mostly in the tip region, was detected, and this material was progressively loosened into the surrounding medium. As a response to potential toxicity, the excess manganese was isolated in vacuoles, which formed an internal barrier against penetration of manganese to the tip area. Elevated manganese concentrations might affect plant reproduction, resulting in anomalies in gamete development. Consequently, the loss in genetic diversity and decreased fruit set ultimately lower yield.

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Lack of visible post-pollination effects in pollen grains of two Dendrobium cultivars: relationship with pollinia ACC, pollen germination, and pollen tube growth

Functional Plant Biology ◽

10.1071/fp07245 ◽

2008 ◽

Vol 35 (2) ◽

pp. 152 ◽

Cited By ~ 3

Author(s):

Kanjana Luangsuwalai ◽

Saichol Ketsa ◽

Apinya Wisutiamonkul ◽

Wouter G. van Doorn

Keyword(s):

Pollen Tube ◽

Ethylene Production ◽

Pollen Tube Growth ◽

Pollen Germination ◽

Flower Colour ◽

Pollen Grains ◽

The Other ◽

Tube Growth ◽

Colour Changes ◽

Acc Content

Dendrobium flowers, pollinated with pollinia from individuals of the same cultivar or other cultivars, usually show rapid post-pollination effects such as floral epinasty, a change in flower colour and early perianth senescence. However, pollination with the pollinia of cv. Karen or cv. Kenny flowers did not produce these effects. We compared these two cultivars with cvv. Pompadour, Willie and Sakura, and tested the hypotheses that the differences were related to levels of 1-aminocyclopropane-1-carboxylic acid (ACC) in the pollinia, ethylene production by the pollinated flower, pollen germination, or pollen tube growth. The pollinia of cvv. Karen and Kenny contained as much ACC as the pollinia of cv. Pompadour, but less than the pollinia of cvv. Willie and Sakura. Ethylene production after pollination with cvv. Karen and Kenny pollinia was much lower than after pollination with pollinia from the other cultivars tested. The pollen grains showed normal germination, but cvv. Karen and Kenny pollen grains exhibited much less tube growth than those of the other cultivars. Pollen tube growth in cv. Pompadour was positively affected by ethylene. Ethylene was required and sufficient for the induction of epinasty, rapid perianth colour changes and early perianth senescence, very similar to the changes after pollination. The absence of these effects after pollination with cvv. Kenny and Karen seems to be due to the low ethylene production induced by the pollinia of these cultivars. This low ethylene production could not be accounted for by the ACC content in the pollinia of cvv. Kenny and Karen.

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Pollen Morphology, Viability, and Germination of Tanacetum vulgare L.

HortScience ◽

10.21273/hortsci.47.3.440 ◽

2012 ◽

Vol 47 (3) ◽

pp. 440-442 ◽

Cited By ~ 1

Author(s):

Nina Devrnja ◽

Jelena Milojević ◽

Ljiljana Tubić ◽

Snežana Zdravković-Korać ◽

Aleksandar Cingel ◽

...

Keyword(s):

Pollen Germination ◽

Pollen Morphology ◽

Average Distance ◽

Pollen Grains ◽

Tube Growth ◽

Tanacetum Vulgare ◽

Medicinal Uses ◽

Spine Length ◽

Pollen Nucleus ◽

Scanning Electron

The pollen morphology of aromatic Tanacetum vulgare L. (Asteraceae), which has wide horticultural and medicinal uses, was examined using light microscopy and scanning electron microscopy. The investigation revealed that pollen grains are radially symmetrical, isopolar, spheroidal, tricolporate, and echinate–perforate. Average pollen length was 21.32 ± 1.12 μm, whereas average pollen width was 20.04 ± 0.98 μm (length:width ratio 0.94). Spine length was 2.72 ± 0.29 μm. Average distance from the spines was 7.15 ± 0.31 μm. Pollen exine consisted of 1–3 pores. Pori are elongated and also with a distinct margin. The porus latitude is smaller than the colpus latitude. One to three perforations were noticed on 1 μm2 of exine. Fluorescein diacetate was used to assess the viability of T. vulgare pollen. The effect of sucrose (1%, 5%, 10%, 20%, and 30%, w/v) on pollen germination and tube growth was evaluated. Overall, the inclusion of sucrose in the medium improved both pollen germination and tube growth. Also, pollen nucleus status was determined. Binucleate and trinucleate mature pollens were observed. Overall, the palynological features of this species may be helpful for further taxonomical and pharmaceutical investigations.

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Metabolomic Analysis of Pollen Grains with Different Germination Abilities from Two Clones of Chinese Fir (Cunninghamia lanceolata (Lamb) Hook)

Molecules ◽

10.3390/molecules23123162 ◽

2018 ◽

Vol 23 (12) ◽

pp. 3162 ◽

Cited By ~ 4

Author(s):

Seif Aldin Dawina Abdallah Fragallah ◽

Pei Wang ◽

Nuo Li ◽

Yu Chen ◽

Sizu Lin

Keyword(s):

Pollen Germination ◽

Early Stage ◽

Pollen Grains ◽

Chinese Fir ◽

Tube Growth ◽

Metabolomic Analysis ◽

Factors Affecting ◽

Germination Process ◽

Germinating Pollen

Pollen grains produce certain metabolites, which can improve or inhibit germination and tube growth. Metabolomic analysis of germinating and growing Chinese fir pollen has not been reported. Therefore, this study aimed to analyse metabolites changes, content and expression in the germinating pollen of Chinese fir. To understand the metabolic differences, two clones from Chinese fir were selected. Metabolomics analyses were performed on three stages (1-, 24- and 48-h) during in vitro pollen germination. The metabolites profiles at different time points were analyzed by using liquid chromatography-mass spectrometry. The results showed that 171 peaks were screened; the corresponding differential metabolites of 121 peaks were classified into nine types of substances. The expression of metabolites showed significant differences across and between clones, and the variation was evident at all germination stages. The expression was obvious at the early stage of germination, which differed clearly from that of the late stage after pollen tube growth. Moreover, the metabolites were mainly enriched in 14 metabolic pathways. Pollen germination and tube growth and metabolites expressions changed per incubation time. Since this work is preliminary, we suggest further investigations to understand the relationship between the differential metabolites and pollen development, and factors affecting pollen germination process.

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PUCHI Regulates Giant Cell Morphology During Root-Knot Nematode Infection in Arabidopsis thaliana

Frontiers in Plant Science ◽

10.3389/fpls.2021.755610 ◽

2021 ◽

Vol 12 ◽

Author(s):

Reira Suzuki ◽

Mizuki Yamada ◽

Takumi Higaki ◽

Mitsuhiro Aida ◽

Minoru Kubo ◽

...

Keyword(s):

Arabidopsis Thaliana ◽

Cell Wall ◽

Genetic Regulation ◽

Three Dimensional ◽

Acid Synthesis ◽

Giant Cells ◽

Loss Of Function ◽

Wild Type ◽

Root Knot Nematode ◽

In The Wild

Parasitic root-knot nematodes transform the host’s vascular cells into permanent feeding giant cells (GCs) to withdraw nutrients from the host plants. GCs are multinucleated metabolically active cells with distinctive cell wall structures; however, the genetic regulation of GC formation is largely unknown. In this study, the functions of the Arabidopsis thaliana transcription factor PUCHI during GC development were investigated. PUCHI expression was shown to be induced in early developing galls, suggesting the importance of the PUCHI gene in gall formation. Despite the puchi mutant not differing significantly from the wild type in nematode invasion and reproduction rates, puchi GC cell walls appeared to be thicker and lobate when compared to the wild type, while the cell membrane sometimes formed invaginations. In three-dimensional (3D) reconstructions of puchi GCs, they appeared to be more irregularly shaped than those in the wild type, with noticeable cell-surface protrusions and folds. Interestingly, the loss-of-function mutant of 3-KETOACYL-COA SYNTHASE 1 showed GC morphology and cell wall defects similar to those of the puchi mutant, suggesting that PUCHI may regulate GC development via very long chain fatty acid synthesis.

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Tapetal 3-Ketoacyl-Coenzyme A Synthases Are Involved in Pollen Coat Lipid Accumulation for Pollen-Stigma Interaction in Arabidopsis

Frontiers in Plant Science ◽

10.3389/fpls.2021.770311 ◽

2021 ◽

Vol 12 ◽

Author(s):

Zaibao Zhang ◽

Huadong Zhan ◽

Jieyang Lu ◽

Shuangxi Xiong ◽

Naiying Yang ◽

...

Keyword(s):

Fatty Acid ◽

Molecular Mechanisms ◽

Coenzyme A ◽

Pollen Grains ◽

Wild Type ◽

Triple Mutant ◽

Long Chain Fatty Acid ◽

Pollen Coat ◽

And Function ◽

Mutant Pollen

Pollen coat lipids form an outer barrier to protect pollen itself and play essential roles in pollen-stigma interaction. However, the precise molecular mechanisms underlying the production, deposition, regulation, and function of pollen coat lipids during anther development remain largely elusive. In lipid metabolism, 3-ketoacyl-coenzyme A synthases (KCS) are involved in fatty acid elongation or very-long-chain fatty acid (VLCFA) synthesis. In this study, we identified six members of the Arabidopsis KCS family expressed in anther. Among them, KCS7, KCS15, and KCS21 were expressed in tapetal cells at anther stages 8–10. Further analysis demonstrated that they act downstream of male sterility 1 (MS1), a regulator of late tapetum development. The kcs7/15/21 triple mutant is fertile. Both cellular observation and lipid staining showed pollen coat lipid was decreased in kcs7/15/21 triple mutant. After landing on stigma, the wild-type pollen grains were hydrated for about 5 min while the kcs7/15/21 triple mutant pollen took about 10 min to hydrate. Pollen tube growth of the triple mutant was also delayed. These results demonstrate that the tapetum-localized KCS proteins are involved in the accumulation of pollen coat lipid and reveal the roles of tapetal-derived pollen coat lipid for pollen-stigma interaction.

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The Tension-sensitive Ion Transport Activity of MSL8 is Critical for its Function in Pollen Hydration and Germination

10.1101/084988 ◽

2016 ◽

Author(s):

Eric S. Hamilton ◽

Elizabeth S. Haswell

Keyword(s):

Pollen Germination ◽

Pollen Viability ◽

Point Mutations ◽

Normal Growth ◽

Loss Of Function ◽

Transport Activity ◽

Wild Type ◽

Gated Channel ◽

In The Wild ◽

Pore Lining

AbstractAll cells respond to osmotic challenges, including those imposed during normal growth and development. Mechanosensitive (MS) ion channels provide a conserved mechanism for regulating osmotic forces by conducting ions in response to increased membrane tension. We previously demonstrated that the MS ion channel MscS-Like 8 (MSL8) is required for pollen to survive multiple osmotic challenges that occur during the normal process of fertilization, and that it can inhibit pollen germination. However, it remained unclear whether these physiological functions required ion flux through a mechanically gated channel provided by MSL8. We introduced two point mutations into the predicted pore-lining domain of MSL8 that disrupted normal channel function in different ways. The Ile711Ser mutation increased the tension threshold of the MSL8 channel while leaving conductance unchanged, and the Phe720Leu mutation severely disrupted the MSL8 channel. Both of these mutations impaired the ability of MSL8 to preserve pollen viability during hydration and to maintain the integrity of the pollen tube when expressed at endogenous levels. When overexpressed in a msl8-4 null background, MSL8I711S could partially rescue loss-of-function phenotypes, while MSL8F720L could not. When overexpressed in the wild type Ler background, MSL8I711S suppressed pollen germination, similar to wild type MSL8. In contrast, MSL8F720L failed to suppress pollen germination and increased pollen bursting, thereby phenocopying the msl8-4 mutant. Thus, an intact MSL8 channel is required to for normal pollen function during hydration and germination. These data establish MSL8 as the first plant MS channel to fulfill previously established criteria for assignment as a mechanotransducer.

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