Tuesday, 20 May 2014 08:31

It's Not Which Lamin but How Much That's Essential

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ABFigure. Expression of lamin-A at various levels in lamin-null
mESCs. (A) A high level of lamin-A (green) forms a smooth
and continuous layer at the periphery of the nucleus
(magenta).(B) When expressed at a low level, lamin-A (green)
forms a discontinuous structure. Scale bar, 5µm.
Once you could pity the lamins. As intermediate filaments, the lamins were often slighted as awkward siblings in between actin and microtubules. Found right under the inner nuclear membrane, lamins were regarded as little more than building materials for the nuclear lamina consisting of additional nuclear proteins. No more. Lamins have come up in the cell world, tied in recent years to transcriptional regulation and linked directly to a rare human developmental disorder of rapid aging called Hutchinson-Gilford progeria syndrome. But their fundamental place in eukaryotic cell biology remained unclear. Lamins are ubiquitously conserved across metazoans but are they essential to cell life?

Now comes a surprising answer from a truly elegant experiment by Yixian Zheng and her colleagues in the Department of Embryology, Carnegie Institution for Science, Baltimore, and collaborators at Northwestern University. As reported recently in Molecular Biology of the Cell (MBoC), Zheng's data say, yes, lamins are essential to nuclear structure but not in the way imagined previously.

Mice and humans have three kinds of lamins—lamin-A/C, lamin-B1, and lamin-B2. How these lamins co-assemble into the nuclear lamina has been unclear, but previous studies imply that one of these lamins might be more critical than the others. The data from Zheng et al. now show that the proper assembly of the nuclear lamina depends on their total concentration. Zheng says "The assembly into a proper structure is not hierarchical. You don't first have to have lamin-B1 and then you recruit lamin-B2 and then you recruit lamin-A/C." This is not to say that certain lamins are not required for the cell to develop further during building of specific tissues, the researchers report. Lamin-A/C is absolutely required to retain the lamina-associated protein emerin, an integral nuclear envelope protein a defect in which is involved in X-linked Emery-Dreifuss muscular dystrophy.

The experiment was possible because the researchers worked with lines of knockout mouse cells each null for a particular lamin or all lamins. Zheng explains that the knockouts gave the researchers a completely null protein background that would not have been possible or easily attainable with an RNAi approach. Combining gene knockout with selective crossbreeding, the researchers were able to generate mouse embryonic stem cells (mESCs) and mouse embryonic fibroblasts with no lamins, a single lamin, or different combinations of two lamins. Lamin concentration was the one variable that guaranteed the smooth organization of lamins within the nuclear lamina. This also ensures even distribution of nuclear pore complexes, the gateways through which all nuclear genetic information must flow to the cytoplasm.

She says that her lab's interest in lamins was encouraged by her collaborator on this MBoC paper, Bob Goldman at Northwestern, who has championed intermediate filament research for years. But Zheng explains that her original decision to look at lamins emerged logically from her longtime concentration on the mitotic spindle. "My lab started to work on lamins from the mitotic spindle angle. The bread and butter projects in my lab have always been mitotic spindle assembly and microtubules. But about eight years ago, we decided to look beyond microtubules and to look at spindle-associated proteins, such as lamins, and their potential influence on spindle morphogenesis."

"In the mitosis field, if a protein affects spindle assembly, the immediate reaction is that it must be essential for cells to exist and propagate because the spindle is essential for cell division." It came as a huge disappointment to Zheng and colleagues when they first learned that mESCs deleted of all lamins survived, divided, and even differentiated into different cell lineages in vitro. To many in the mitosis field, these results seemed to seal the fate of lamins as not having a role in spindle function. "But that is a wrong interpretation because mitotic spindle is not just for cell division. It's also required for the orientation of cell division and for proper separation of cell fate determinants. Our recent studies in the neural progenitor cells in mice indeed show that B-type lamins are required for spindle orientation," Zheng says.

The multiple interactions lamins make in interphase and mitotic cells have made it difficult to pinpoint how these proteins work in the spindle in mitosis and in the nucleus in interphase. Zheng says, "One way we may be able to make progress is to study lamins in the context of specific cell and tissue types in the context of development and homeostasis."

The rigid separation of cell biology into subfields where some people worked only on cytoskeleton while others toiled inside the nucleus made sense in earlier days, says Zheng, but not any longer. "Cell biology for many years has taken a reductionist approach in order to understand complex processes such as the assembly of spindle and nucleus in tissue culture cells." But now, she says, "It's time to understand how cytoskeleton and nucleus talk to one another to make the cell work not just in tissue culture cells but also in more physiologically relevant contexts."

"The creation of lamin knockout cells and mice has opened the door to address basic questions such as whether different lamins have unique roles in their assembly into a smooth network of polymers in the nuclear lamina in interphase and how lamins influence the spindle in mitosis," Zheng says. The payoff is clear because understanding the shared and unique functions of different lamins is a critical first step to further decipher their functions in the nucleus and spindle."

"The role of nuclear lamins in genome organization is an emerging field because people are really starting to see that understanding the connection between nuclear lamins and the cytoskeleton could shed light on development and tissue building," she says. "Hopefully, nuclear lamina will help us to understand the morphological foundation for nuclear organization and transcription. At least, that's my dream."

Guo Y, Kim Y, Shimi T, Goldman RD, Zheng Y (2014). Concentration-dependent lamin assembly and its roles in the localization of other nuclear proteins. Mol Biol Cell 25, 1287–1297. http://www.molbiolcell.org/content/25/8/1287.abstract

John Fleischman

John is ASCB Senior Science Writer and the author among other things of two nonfiction books for older children, "Phineas Gage: A Gruesome But True Story About Brain Science" and "Black & White Airmen," both from Houghton-Mifflin-Harcourt, Boston.

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