SYMPOSIA / S15-5 : DEVELOPMENT AND SIGNALLING IN ARABIDOPSIS THALIANA POLLEN

(David Twell) 한국생화학분자생물학회 (구 한국생화학회) 1999 6th IUBMB SEOUL CONFERENCE Vol.- No.-

Arabidopsis fused kinase TWO-IN-ONE dominently inhibits male meiotic cytokinesis

Sung Aeong Oh,David Twell,Soon Ki Park 한국육종학회 2013 한국육종학회 심포지엄 Vol.2013 No.07

Arabidopsis Fused kinase TWO-IN-ONE (TIO) controls phragmoplast expansion and interacts with the Kinesin-12 subfamily proteins that anchor the plus ends of interdigitating microtubules (MTs) in the phragmoplast midzone. Previous analyses of loss-of-function mutants and RNA interference lines revealed that TIO positively controls both somatic and gametophytic cell cytokinesis, however, knowledge of the full spectrum of TIO functions during plant development remains incomplete. In order to further characterize TIO functions, we expressed TIO and a range of TIO variants under control of its own promoter in wild type Arabidopsis plants. We discovered that TIO-overexpressing transgenic lines produce enlarged pollen grains, arising from incomplete cytokinesis during male meiosis, and showed sporophytic abnormalities indicating polyploidy. These phenotypes arose independently in TIO variants that abolished either gametophytic function or the ability of TIO to interact with Kinesin-12 subfamily proteins. Interaction assays in yeast showed TIO to bind to AtNACK2/TETRASPORE and plants doubly homozygous for kinesin-12a and kinesin-12b knockout mutations to produce enlarged pollen grains. Our results show that TIO dominantly inhibits male meiotic cytokinesis in a dosage dependent manner that may involve direct binding to acomponent of the canonical NACK-PQR cytokinesis signaling pathway.

The <i>SIDECAR POLLEN</i> gene encodes a microspore‐specific LOB/AS2 domain protein required for the correct timing and orientation of asymmetric cell division

Oh, Sung Aeong,Park, Keun Sang,Twell, David,Park, Soon Ki Blackwell Publishing Ltd 2010 The Plant journal Vol.64 No.5

<P><B>Summary</B></P><P>Cellular patterning and differentiation in plants depend on the balance of asymmetric and symmetric divisions. Patterning of the male gametophyte (pollen grains) in flowering plants requires asymmetric division of the microspore followed by a symmetric division of the germ cell to produce three highly differentiated cells: a single vegetative cell and two sperm cells. In Arabidopsis <I>sidecar pollen</I> (<I>scp</I>) mutants a proportion of microspores first divide symmetrically, and then go on to produce ‘four‐celled’ pollen with an extra vegetative cell; however, details of the timing and origin of phenotypic defects in <I>scp</I> and the identity of the <I>SCP</I> gene have remained obscure. Comparative analysis of the original hypomorphic <I>scp‐1</I> allele and a T‐DNA‐induced null allele, <I>scp‐2</I>, revealed that in the absence of SCP, microspores undergo normal nuclear positioning, but show delayed entry into mitosis, increased cell expansion and alterations in the orientation of nuclear division. We identified the <I>SCP</I> gene to encode a male gametophyte‐specific LATERAL ORGAN BOUNDARIES DOMAIN/ASYMMETRIC LEAVES 2‐like (LBD/ASL) protein that is expressed in microspore nuclei in a tightly regulated phase‐specific manner. Therefore, our study demonstrates that the correct patterning of male gametophyte depends on the activity of a nuclear LBD/ASL family protein that is essential for the correct timing and orientation of asymmetric microspore division.</P>

Overexpression of TWO-IN-ONE Domains Inhibits Cytokinesis in Arabidopsis

Oh Sung-Aeong,Lee Sang Ju,김정회,Twell David,박순기 한국식물학회 2022 Journal of Plant Biology Vol.65 No.4

Plant cells undergoing cytokinesis build a cell plate between daughter nuclei and expand it centrifugally toward the parental plasma membrane. This process is enabled by the phragmoplast, a plant-specific and highly dynamic microtubule (MT)-based array. A signaling module required for phragmoplast expansion consisting of TWO-IN-ONE (TIO) and two Kinesin-12 members is described in Arabidopsis thaliana. TIO is a deeply conserved Fused kinase that has a divergent plant-specific role in cytokinesis. TIO has two conserved domains, the N-terminal kinase and the C-terminal ARM/HEAT domains, which are crucial for function and interaction with the phragmoplast MT plus-end associated Kinesin-12A and -12B. Here, we further explored the importance of TIO domains for cytokinesis in an overexpression approach. In addition, the functionality of a putative activation loop in the kinase domain was validated by substituting two invariant threonine and serine residues. Phenotypic analysis reveals that overexpression of the domains severely interferes with cytokinesis in pollen and in somatic cells. Furthermore, measuring the level of rescue for gametophytic cytokinesis in tio-3 null mutant background reveals that the putative kinase activation loop is required for TIO function and likely operates via reversible phosphorylations. Therefore, this study enhances understanding of TIO signaling and domain functions in cytokinesis.

Map-based cloning and functional anlysis of sidecar pollen, Arabidopsis gametophytic mutant

Keun Sang Park,Sung Aeong Oh,Hyun Kyung Bae,David Twell,Soon Ki Park 한국육종학회 2007 한국육종학회 학술발표회 발표요지 Vol.39 No.1

The tobacco MAP215/Dis1-family protein TMBP200 is required for the functional organization of microtubule arrays during male germline establishment

Oh, Sung Aeong,Pal, Madhumita Das,Park, Soon Ki,Johnson, James Andrew,Twell, David Oxford University Press 2010 Journal of experimental botany Vol.61 No.4

<P>The haploid microspore division during pollen development in flowering plants is an intrinsically asymmetric division which establishes the male germline for sexual reproduction. Arabidopsis <I>gem1</I> mutants lack the male germline as a result of disturbed microspore polarity, division asymmetry, and cytokinesis and represent loss-of-function mutants in <I>MOR1/GEM1,</I> a plant orthologue of the conserved MAP215/Dis1 microtubule associated protein (MAP) family. This provides genetic evidence for the role of MAP215/Dis1 in the organization of gametophytic microtubule arrays, but it has remained unknown how microtubule arrays are affected in <I>gem1</I> mutant microspores. Here, novel male gametophytic microtubule-reporter <I>Nicotiana tabacum</I> plants were constructed, expressing a green fluorescent protein-α-TUBULIN fusion protein (GFP-TUA6) under the control of a microspore-specific promoter. These plants allow effective visualization of all major male gametophytic microtubule arrays and provide useful tools to study the regulation of microtubule arrays by MAPs and other effectors. Depletion of <I>TMBP200</I>, a tobacco homologue of <I>MOR1/GEM1</I> in gametophytic microtubule-reporter plants using microspore-targeted RNA interference, induced defects in microspore polarity, division asymmetry and cytokinesis that were associated with striking defects in phragmoplast position, orientation, and structure. Our observations further reveal a requirement for <I>TMBP200</I> in gametophytic spindle organization and a novel role in spindle position and orientation in polarized microspores. These results provide direct evidence for the function of MAP215/Dis1 family protein TMBP200 in the organization of microtubule arrays critical for male germline formation in plants.</P>

Arabidopsis Fused kinase TWO-IN-ONE dominantly inhibits male meiotic cytokinesis

Oh, Sung Aeong,Bourdon, Valé,rie,Dickinson, Hugh G.,Twell, David,Park, Soon Ki Springer-Verlag 2014 Plant reproduction Vol.27 No.1

The gemini pollen 3 gene is required for nuclear division of pollen meiosis and mitosis in Arabidopsis

Ji-Eun Jeon,Sung Aeong Oh,Hyo-Jin Park,Gyun Jang Kim,Keun Sang Park,David Twell,Jong Tae Song,Moon-Soo Soh,Soon Ki Park 한국육종학회 2014 한국육종학회 심포지엄 Vol.2014 No.07

The correct development of male gametophytes (pollen grains) in flowering plants is essential for proliferate in gamete production. Here we have taken a map-based cloning approach using Arabidopsis male gametophytic mutant, named gemini pollen3 (gem3) to identify and characterize key gene that is expressed gametophytically for the completion of microgametogenesis focusing on genes which control cell division and cell fate determination. Previously reported gem1 and gem2 mutants with similar characteristics to gem3 that are disturbed at asymmetric division and cytokinesis at pollen mitosis I (PMI) in Arabidopsis. However, gem3 was mapped to a different genetic locus, and pollen developmental analysis revealed that gem3 exert an effect at meiosis and mitosis causing complete sterility. We also discovered that gem3 homozygous lines produce aberrant pollen grains, arising from incomplete cytokinesis during male meiosis with sporophytic phenotypes of twisted-shape leaves, large flowers. This mutation shows reduced genetic transmission of gem3 allele through male gametophyte. In previous results, the gem3 locus was confirmed by mapping to the region located on chromosome 5. To further confirm strong candidate gene, we performed sequencing and genetic complementation analysis. Currently, we are performing functional studies of the gem3 gene for the better understanding of molecular mechanisms that control asymmetric division at meiosis and mitosis during pollen development.

Nucleoporin MOS7/Nup88 is required for mitosis in gametogenesis and seed development in <i>Arabidopsis</i>

Park, Guen Tae,Frost, Jennifer M.,Park, Jin-Sup,Kim, Tae Ho,Lee, Jong Seob,Oh, Sung Aeong,Twell, David,Brooks, Janie Sue,Fischer, Robert L.,Choi, Yeonhee National Academy of Sciences 2014 PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF Vol.111 No.51

<P><B>Significance</B></P><P>During plant reproduction, meiosis generates haploid spores that undergo mitoses, forming gametophytes, in male and female parts of the flower. Haploid cells within gametophytes differentiate into sperm and eggs, which form the next generation. We discovered that the <I>MOS7</I> (Modifier of Snc1,7) gene plays a critical role in microtubule organization and dynamics during mitosis in <I>Arabidopsis</I> sexual reproduction. MOS7 absence results in impaired gametogenesis, failure of ovule and pollen development, and seed abortion. Although mitoses during gamete formation are unique to plants, the regulation of mitosis itself is highly conserved between plants and vertebrates. Notably, we find that MOS7 protein primarily produced prior to meiosis is inherited by gametophytes, and, only if there is sufficient MOS7 protein, will mitosis correctly occur during gametogenesis.</P><P>Angiosperm reproduction is characterized by alternate diploid sporophytic and haploid gametophytic generations. Gametogenesis shares similarities with that of animals except for the formation of the gametophyte, whereby haploid cells undergo several rounds of postmeiotic mitosis to form gametes and the accessory cells required for successful reproduction. The mechanisms regulating gametophyte development in angiosperms are incompletely understood. Here, we show that the nucleoporin Nup88-homolog MOS7 (Modifier of Snc1,7) plays a crucial role in mitosis during both male and female gametophyte formation in <I>Arabidopsis thaliana</I>. Using a mutagenesis screen, we identify the <I>mos7-5</I> mutant allele, which causes ovule and pollen abortion in <I>MOS7/mos7-5</I> heterozygous plants, and preglobular stage embryonic lethality in homozygous <I>mos7-5</I> seeds. During interphase, we show that MOS7 is localized to the nuclear membrane but, like many nucleoporins, is associated with the spindle apparatus during mitosis. We detect interactions between MOS7 and several nucleoporins known to control spindle dynamics, and find that in pollen from <I>MOS7</I>/<I>mos7-5</I> heterozygotes, abortion is accompanied by a failure of spindle formation, cell fate specification, and phragmoplast activity. Most intriguingly, we show that following gamete formation by <I>MOS7/mos7-5</I> heterozygous spores, inheritance of either the <I>MOS7</I> or the <I>mos7-5</I> allele by a given gamete does not correlate with its respective survival or abortion. Instead, we suggest a model whereby MOS7, which is highly expressed in the Pollen- and Megaspore Mother Cells, enacts a dosage-limiting effect on the gametes to enable their progression through subsequent mitoses.</P>

Arabidopsis Fused kinase and the Kinesin‐12 subfamily constitute a signalling module required for phragmoplast expansion

Oh, Sung Aeong,Allen, Trudie,Kim, Gyun Jang,Sidorova, Anna,Borg, Michael,Park, Soon Ki,Twell, David Blackwell Publishing Ltd 2012 The Plant journal Vol.72 No.2

<P><B>Summary</B></P><P>The conserved Fused kinase plays vital but divergent roles in many organisms from Hedgehog signalling in <I>Drosophila</I> to polarization and chemotaxis in <I>Dictyostelium.</I> Previously we have shown that Arabidopsis Fused kinase termed TWO‐IN‐ONE (TIO) is essential for cytokinesis in both sporophytic and gametophytic cell types. Here using <I>in vivo</I> imaging of GFP‐tagged microtubules in dividing microspores we show that TIO is required for expansion of the phragmoplast. We identify the phragmoplast‐associated kinesins, PAKRP1/Kinesin‐12A and PAKRP1L/Kinesin‐12B, as TIO‐interacting proteins and determine TIO‐Kinesin‐12 interaction domains and their requirement in male gametophytic cytokinesis. Our results support the role of TIO as a functional protein kinase that interacts with Kinesin‐12 subfamily members mainly through the C‐terminal ARM repeat domain, but with a contribution from the N‐terminal kinase domain. The interaction of TIO with Kinesin proteins and the functional requirement of their interaction domains support the operation of a Fused kinase signalling module in phragmoplast expansion that depends upon conserved structural features in diverse Fused kinases.</P>