Cellular and Molecular Life Sciences

, Volume 75, Issue 9, pp 1687–1706 | Cite as

The ancient claudin Dni2 facilitates yeast cell fusion by compartmentalizing Dni1 into a membrane subdomain

  • M.-Ángeles Curto
  • Sandra Moro
  • Francisco Yanguas
  • Carmen Gutiérrez-González
  • M.-Henar Valdivieso
Original Article


Dni1 and Dni2 facilitate cell fusion during mating. Here, we show that these proteins are interdependent for their localization in a plasma membrane subdomain, which we have termed the mating fusion domain. Dni1 compartmentation in the domain is required for cell fusion. The contribution of actin, sterol-dependent membrane organization, and Dni2 to this compartmentation was analysed, and the results showed that Dni2 plays the most relevant role in the process. In turn, the Dni2 exit from the endoplasmic reticulum depends on Dni1. These proteins share the presence of a cysteine motif in their first extracellular loop related to the claudin GLWxxC(8–10 aa)C signature motif. Structure–function analyses show that mutating each Dni1 conserved cysteine has mild effects, and that only simultaneous elimination of several cysteines leads to a mating defect. On the contrary, eliminating each single cysteine and the C-terminal tail in Dni2 abrogates Dni1 compartmentation and cell fusion. Sequence alignments show that claudin trans-membrane helixes bear small-XXX-small motifs at conserved positions. The fourth Dni2 trans-membrane helix tends to form homo-oligomers in Escherichia plasma membrane, and two concatenated small-XXX-small motifs are required for efficient oligomerization and for Dni2 export from the yeast endoplasmic reticulum. Together, our results strongly suggest that Dni2 is an ancient claudin that blocks Dni1 diffusion from the intercellular region where two plasma membranes are in close proximity, and that this function is required for Dni1 to facilitate cell fusion.


Actin Mating Membrane microdomains Small-XXX-small Sterols Tight junction Schizosaccharomyces pombe 



Chloramphenicol acetyl transferase


Extra-cellular loop


Endoplasmic reticulum


Green fluorescent protein


Membrane fusion domain


Post-synaptic density protein PSD95, Drosophila disc large tumour suppressor Dlg1, and zonula occludens-1 zo-1 proteins


Tight junction


Trans-membrane Domain





We thank E. Keck for the English revision. We are indebted to S. Moreno, D. Mulvihill, O. Nielsen, P. Nurse, P. Pérez, J.C. Ribas, Y. Sanchez, C. Shimoda, and K. Takegawa for strains and plasmids. A. Senes and C. Armstrong are acknowledged for providing plasmids and the guidance to perform the TOXCAT analyses. Financial support from the Ministerio de Economía y Competitividad (MINECO, Spain)/European Union FEDER program (BFU2013-48582-C2-2-P) and Junta de Castilla y León (Grant SA073U14) made this work possible. MAC, SM, and FY were supported by FPU fellowships from the Ministerio de Educación. CGG was supported by the Sistema de Garantía Juvenil program from the Ministerio de Empleo y Seguridad Social, Spain.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.

Supplementary material

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Supplementary material 8 (PDF 124 kb)


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Authors and Affiliations

  1. 1.Departamento de Microbiología y GenéticaUniversidad de SalamancaSalamancaSpain
  2. 2.Instituto de Biología Funcional y Genómica (IBFG), Consejo Superior de Investigaciones Científicas (CSIC)SalamancaSpain

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