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Research

 

Why polarity?

Cell polarity is essential for the proper functioning of many cell types, because the cell function is optimized by

the cell shape, and the cell shape is determined by polarity. Filamentous fungi grow by tip-extension. The mode

of growth depends on stable polarity at the hyphal tip. How is the polarity regulated in filamentous fungi?

Why do filamentous fungi grow filamentous?

 

Cell end marker

We found landmark proteins named “cell end markers”, which control the polarity. The cell end markers, TeaA

and TeaR, localize at hyphal tips interdependently. TeaA is delivered to hyphal tips by growing microtubules

(MTs), anchored to the hyphal tip cortex though the interaction with TeaR, and it indirectly interacts with the

formin SepA which forms actin cables that are required for secretion vesicle transport and thus for polarized

growth. The teaA and teaR deletion mutants showed defects in polarity maintenance, which lead to curved or

zig-zag growing hyphae. Thus TeaA transmits positional information regulated by MTs to the actin cytoskeleton

at hyphal tips. The mechanism is required for the polarity maintenance and the growth direction of hyphae.

 




MTs and plasma membrane

The interaction between MT plus end and the plasma membrane is important for the establishment and

maintenance of polarity. MT plus-end localizing proteins (+TIPs) regulate MT plus-end dynamics. AlpA, a

XMAP215 family protein and one of +TIPs, promotes MT growth and functions as MT polymerase. The functional

connection between the cell end marker TeaA at the plasma membrane and AlpA at MT plus-ends is important

for the proper regulation of MT growth at hyphal tips. In this way, we focus on the connection among MT plus

ends, plasma membrane and actin cytoskeleton for polarized growth.

 

 

 

 


Visualization of membrane domains by super-resolution microscopy

Apical sterol-rich membrane domains (SRDs) are gaining attention for their important roles in polarized growth

of filamentous fungi, however their exact figure, roles and formation mechanisms remain rather unclear. We

selected genes involved in membrane recycle, raft formation and lipid transport, etc. We are investigating their roles of

on the formation and maintenance of SRD and their interplay with the cytoskeletons.

     The SRDs are revealed to be a mixture of lipid raft and non-raft microdomains, however the size of rafts is

ranging between 10 and 200 nm and is thus too small to detect by conventional light microscopy which has a

resolution limit of 250 nm. In recent years, super-resolution microscope techniques have been improving and

breaking the diffraction limit of conventional light microscopy. One of the techniques is photoactivation

localization microscopy (PALM) using photoswitchable (or photoactivatable) fluorophores. The lateral image

resolution as high as 20 nm which can be acquired via this method will be a powerful tool to investigate the

relation of lipid membrane domains and protein localization in living cells deeply. Size, number, distribution and

dynamics of membrane domains, and dynamics of single molecules are subjects of our research. Additionally 

we are investing the MT and actin cytoskeleton, and their relation with membrane domains. 

 



 

 

List of publications (corresponding author * )

25.        Takeshita N*. Coordinated process of polarized growth in filamentous fungi.

            Biosci. Biotechnol. Biochem. (2016) Review

Special issue; Fungal Molecular Biology and Biotechnology

 

24.        Bergs A, Ishitsuka Y, Evangelinos M, Nienhaus GU, Takeshita N*. 

            Dynamics of actin cables in polarized growth of the filamentous fungus 

Aspergillus nidulans. Frontiers in Microbiology (2016) 7:682.

            http://journal.frontiersin.org/article/10.3389/fmicb.2016.00682/full

 

23.        Ishitsuka Y, Savage N, Li Y, Bergs A, Grün N, Kohler D, Donnelly R, Nienhaus GU,

            Fischer R, Takeshita N*.

            Super-resolution microscopy reveals a dynamic picture of cell polarity maintenance

            during directional growth. Science Advances (2015) 1, e1500947.

            http://advances.sciencemag.org/content/1/10/e1500947

 

22.        Manck R, Ishitsuka Y, Herrero S, Takeshita N, Nienhaus GU, Fischer R.

            Genetic evidence for a microtubule-capture mechanism during polar growth of

            Aspergillus nidulans. J Cell Sci. (2015) 128, 3569-82.

 

21.        Bühler N, Hagiwara D, Takeshita N*. Functional analysis of sterol transporter

            orthologues (OSHs) in the filamentous fungus Aspergillus nidulans

            Eukaryot Cell (2015) 14, 908-21.

 

20.        Takeshita N*, Wernet V, Tsuizaki M, Grün N, Hoshi HO, Ohta A, Fischer R, Horiuchi H.

            Transportation of Aspergillus nidulans class III and V chitin synthases to the hyphal

            tips depends on conventional kinesin. PLoS One (2015) 10(5):e0125937.

 

19.        Baumann S, Takeshita N, Grün N, Fischer R, Feldbrügge M.

            Live cell imaging of endosomal trafficking in fungi. 

            Meth. Mol. Biol. (2015) 1270,347-63.

 

18.        Pöhlmann J, Risse C, Seidel C, Pohlmann T, Jakopec V, Walla E, Ramrath P, 

Takeshita N, Feldbrügge M, Fischer R, Fleig U.

            The Vip1 inositol polyphosphate kinase family regulates polarized growth and modulates

the microtubule cytoskeleton in fungi. PLoS Genet. (2014) 10(9) e1004586.

 

17.        Herrero S, Takeshita N, Fischer R.

            The F-box protein RcyA controls the turnover of the kinesin-7 motor KipA

in Aspergillus nidulans. Eukaryot. Cell (2014) 18, 1085-94. 

 

16.        Takeshita N*, Manck R, Grün N, de Vega S, Fischer R.

            Interdependence of the actin and the microtubule cytoskeleton during fungal growth.

            Curr. Opin. Microbiol. (2014) 20C:34-41. Review

 

15.        Takeshita N*, Mania D, Herrero S, Ishitsuka Y, Nienhaus GU, Podolski M, 

Howard J, Fischer R. The cell end marker TeaA and the microtubule polymerase AlpA

contribute to microtubule guidance at the hyphal tip cortex of Aspergillus nidulans for

polarity maintenance. J. Cell Sci. (2013) 126, 5400-11.

 

14.     Takeshita N*, Diallinas G, Fischer R. The role of Flotillin FloA in the formation of apical

sterol-rich membrane domains (SRDs) and polarity determination in the filamentous

fungus Aspergillus nidulans. Mol. Microbiol. (2012) 83, 1136-52.

  

13.     Takeshita N*, Fischer R. On the role of microtubules, cell end markers, and septal

         microtubule organizing centres on site selection for polar growth in Aspergillus nidulans.

         Fungal. Biol. (2011) 115, 506-517. The 200th Anniversary of the hyphae, special issue

  

12.    Herrero S, Takeshita N, Fischer R. The Aspergillus nidulans CENP-E kinesin motor KipA

         interacts with the fungal homolog of the centromere-assicated protein CENP-H at the

         kinetochore. Mol. Microbiol. (2011) 80, 981-994.

  

11.     Mania D, Hilpert K, Ruden S, Fischer R, Takeshita N*.

         Screening for antifungal peptides and their modes of action in Aspergillus nidulans.

         Applied Environ. MIcrobiol. (2010) 76, 7102-7108.

 

10.     Harris SD, Turner G, Meyer V, Espeso EA, Specht T, Takeshita N, Helmstedt K.

          Morphology and development in Aspergillus nidulans: a complex puzzle.

          Fungal Genet. Biol. (2009) 46 Suppl 1:S82-S92.

 

9.       Higashitsuji Y, Herrero S, Takeshita N, Fischer R.

         The cell end marker protein TeaC is involved in growth directionality and septation

         in Aspergillus nidulans. Eukaryot. Cell (2009) 8, 957-67.

 

8.       Tsuizaki M, Takeshita N, Ohta A, Horiuchi H. Myosin motor-like domain of the class VI

         chitin synthase CsmB is essential to its functions in Aspergillus nidulans.

         Biosci. Biotechnol. Biochem. (2009) 73, 1163-7.

 

7.        Wortman JR, Gilsenan JM, Joardar V, Deegan J, Clutterbuck J, Andersen MR, Archer D, Bencina M, Braus G, Coutinho P, von Döhren H, Doonan J, Driessen AJ, Durek P, Espeso E, Fekete E, Flipphi M, Estrada CG, Geysens S, Goldman G, de Groot PW, Hansen K, Harris SD, Heinekamp T, Helmstaedt K, Henrissat B, Hofmann G, Homan T, Horio T, Horiuchi H, James S, Jones M, Karaffa L, Karányi Z, Kato M, Keller N, Kelly DE, Kiel JA, Kim JM, van der Klei IJ, Klis FM, Kovalchuk A, Krasevec N, Kubicek CP, Liu B, Maccabe A, Meyer V, Mirabito P, Miskei M, Mos M, Mullins J, Nelson DR, Nielsen J, Oakley BR, Osmani SA, Pakula T, Paszewski A, Paulsen I, Pilsyk S, Pócsi I, Punt PJ, Ram AF, Ren Q, Robellet X, Robson G, Seiboth B, van Solingen P, Specht T, Sun J, Taheri-Talesh N, Takeshita N, Ussery D, vanKuyk PA, Visser H, van de Vondervoort PJ, de Vries RP, Walton J, Xiang X, Xiong Y, Zeng AP, Brandt BW, Cornell MJ, van den Hondel CA, Visser J, Oliver SG, Turner G.

         The 2008 update of the Aspergillus nidulans genome annotation: a community effort.

         Fungal Genet. Biol. (2009) 46 Suppl 1:S2-13. 

 

6.       Fischer R, Zekert N, Takeshita N*.

Polarized growth in fungi – interplay among the cytoskeleton, positional markers, and

membrane domains. Mol. Microbiol. (2008) 68, 813-826. Review

 

5.       Takeshita N, Higashitsuji Y, Konzack S, Fischer R.

         Apical sterol-rich membranes are essential for localizing cell end markers that determine

 growth directionality in the filamentous fungus Aspergillus nidulans.

         Mol. Biol.Cell (2008) 19, 339-351.

  

4.       Takeshita N, Vienken K, Rolbetzki A, Fischer R.

         The Aspergillus nidulans putative kinase, KfsA (kinase for septation), plays a role in

         septation and is required for efficient asexual spore formation.

         Fungal Genet. Biol. (2007) 44, 1205-1214.

  

3.       Takeshita N, Yamashita S, Ohta A, Horiuchi H. Aspergillus nidulans class V and VI chitin

         synthases with a myosin motor-like domain, CsmA and CsmB, perform compensatory

         functions that are essential for hyphal tip growth. Mol. Microbiol. (2006) 59, 1380-1349.

 

2.       Takeshita N, Ohta A, Horiuchi H. CsmA, a class V chitin synthase with a myosin motor-

         like domain, is localized through direct interaction with the actin cytoskeleton

         in Aspergillus nidulans. Mol. Biol. Cell (2005) 16, 1961-1970.

 

1.       Takeshita N, Ohta A, Horiuchi H. csmA, a gene encoding a class V chitin synthase with

a myosin motor-like domain of Aspergillus nidulans, is translated as a single polypeptide

and regulated in response to osmotic conditions.

         Biochem. Biophys. Res. Commun. (2002) 298, 103-109.

 

Book

2.         Etxebeste O, Takeshita N. (2015) Advanced microscopy methods for the study of

            protein localization, interaction and dynamics in filamentous fungi. 

            Springer. Advanced Microscopy in Mycology.

 

1.         Fischer R, Takeshita N, Doonan J. (2007)

            Cytoskeleton, Polarized Growth, and the Cell Cycle in Aspergillus nidulans.

            The Aspegilli: Genomics, Medical Applications, Biotechnology, and Research Methods.