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Tts1, the fission yeast homologue of the TMEM33 family, functions in NE remodeling during mitosis.

Zhang D, Oliferenko S - Mol. Biol. Cell (2014)

Bottom Line: An amphipathic helix located at the C-terminus of Tts1 is important for ER shaping and modulating the mitotic NPC distribution.Of interest, the evolutionarily conserved residues at the luminal interface of the third transmembrane region function specifically in promoting SPB-NE insertion.Our data illuminate cellular requirements for remodeling the NE during "closed" nuclear division and provide insight into the structure and functions of the eukaryotic TMEM33 family.

View Article: PubMed Central - PubMed

Affiliation: Temasek Life Sciences Laboratory, National University of Singapore, Singapore 117604 zhangdan@tll.org.sg snezhana.oliferenko@kcl.ac.uk.

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The mitotic SPBs are compromised in recruiting the γ-tubulin complex in cut11-6 tts1Δ cells. (A) Time-lapse maximum z-projection images of spinning disk confocal stacks of cells with indicated genetic backgrounds coexpressing GFP-Atb2, Pcp1-mCherry, and the nucleoplasm marker protein GST-NLS-mCherry. An aster-like short spindle in early mitotic tts1Δ cells is denoted by an arrow. (B, C) Time-lapse maximum z-projection images of spinning disk confocal stacks of cells with indicated genetic backgrounds coexpressing Pcp1-mCherry and Alp4-GFP (B) or Alp6-GFP (C). Cells were grown and imaged at 24°C, the permissive temperature for cut11-6 mutant. The elapsed time is shown in minutes. Scale bars, 5 μm.
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Figure 5: The mitotic SPBs are compromised in recruiting the γ-tubulin complex in cut11-6 tts1Δ cells. (A) Time-lapse maximum z-projection images of spinning disk confocal stacks of cells with indicated genetic backgrounds coexpressing GFP-Atb2, Pcp1-mCherry, and the nucleoplasm marker protein GST-NLS-mCherry. An aster-like short spindle in early mitotic tts1Δ cells is denoted by an arrow. (B, C) Time-lapse maximum z-projection images of spinning disk confocal stacks of cells with indicated genetic backgrounds coexpressing Pcp1-mCherry and Alp4-GFP (B) or Alp6-GFP (C). Cells were grown and imaged at 24°C, the permissive temperature for cut11-6 mutant. The elapsed time is shown in minutes. Scale bars, 5 μm.

Mentions: To examine mitotic spindle formation carefully, we performed time-lapse analyses of cells expressing fluorescent protein markers for the spindle (GFP-Atb2), the SPBs (Pcp1-mCherry), and the nucleoplasm (GST-NLS-mCherry) at 24°C. Wild type cells initiated bipolar spindle assembly within 3 min (2.5 ± 0.8 min, n = 11) after depolymerization of the interphase microtubule arrays (Figure 5A). tts1Δ cells assembled short spindles 2.8 ± 2.0 min (n = 18) after the depolymerization of cytoplasmic microtubules. However, in line with our immunofluorescence data, some tts1Δ cells went through a short but noticeable stage with microtubules forming small, aster-like structures (seven of 18 cells, indicated by an arrow in Figure 5A). All tts1Δ cells subsequently assembled bipolar spindles and completed mitosis.


Tts1, the fission yeast homologue of the TMEM33 family, functions in NE remodeling during mitosis.

Zhang D, Oliferenko S - Mol. Biol. Cell (2014)

The mitotic SPBs are compromised in recruiting the γ-tubulin complex in cut11-6 tts1Δ cells. (A) Time-lapse maximum z-projection images of spinning disk confocal stacks of cells with indicated genetic backgrounds coexpressing GFP-Atb2, Pcp1-mCherry, and the nucleoplasm marker protein GST-NLS-mCherry. An aster-like short spindle in early mitotic tts1Δ cells is denoted by an arrow. (B, C) Time-lapse maximum z-projection images of spinning disk confocal stacks of cells with indicated genetic backgrounds coexpressing Pcp1-mCherry and Alp4-GFP (B) or Alp6-GFP (C). Cells were grown and imaged at 24°C, the permissive temperature for cut11-6 mutant. The elapsed time is shown in minutes. Scale bars, 5 μm.
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Related In: Results  -  Collection

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Figure 5: The mitotic SPBs are compromised in recruiting the γ-tubulin complex in cut11-6 tts1Δ cells. (A) Time-lapse maximum z-projection images of spinning disk confocal stacks of cells with indicated genetic backgrounds coexpressing GFP-Atb2, Pcp1-mCherry, and the nucleoplasm marker protein GST-NLS-mCherry. An aster-like short spindle in early mitotic tts1Δ cells is denoted by an arrow. (B, C) Time-lapse maximum z-projection images of spinning disk confocal stacks of cells with indicated genetic backgrounds coexpressing Pcp1-mCherry and Alp4-GFP (B) or Alp6-GFP (C). Cells were grown and imaged at 24°C, the permissive temperature for cut11-6 mutant. The elapsed time is shown in minutes. Scale bars, 5 μm.
Mentions: To examine mitotic spindle formation carefully, we performed time-lapse analyses of cells expressing fluorescent protein markers for the spindle (GFP-Atb2), the SPBs (Pcp1-mCherry), and the nucleoplasm (GST-NLS-mCherry) at 24°C. Wild type cells initiated bipolar spindle assembly within 3 min (2.5 ± 0.8 min, n = 11) after depolymerization of the interphase microtubule arrays (Figure 5A). tts1Δ cells assembled short spindles 2.8 ± 2.0 min (n = 18) after the depolymerization of cytoplasmic microtubules. However, in line with our immunofluorescence data, some tts1Δ cells went through a short but noticeable stage with microtubules forming small, aster-like structures (seven of 18 cells, indicated by an arrow in Figure 5A). All tts1Δ cells subsequently assembled bipolar spindles and completed mitosis.

Bottom Line: An amphipathic helix located at the C-terminus of Tts1 is important for ER shaping and modulating the mitotic NPC distribution.Of interest, the evolutionarily conserved residues at the luminal interface of the third transmembrane region function specifically in promoting SPB-NE insertion.Our data illuminate cellular requirements for remodeling the NE during "closed" nuclear division and provide insight into the structure and functions of the eukaryotic TMEM33 family.

View Article: PubMed Central - PubMed

Affiliation: Temasek Life Sciences Laboratory, National University of Singapore, Singapore 117604 zhangdan@tll.org.sg snezhana.oliferenko@kcl.ac.uk.

Show MeSH