April 2, 2014

Nuclear envelope breakdown is far prettier than my own breakdown when I realized that Girl Scout “cookie season” is over. Today’s image is from a paper that describes the importance of SUN proteins in nuclear envelope breakdown.

Early in mitosis, a cell’s nuclear envelope breaks down to allow the attachment of chromosomes to the mitotic spindle. Nuclear envelope breakdown (NEBD) depends on a tearing process, during which microtubules pull the nuclear envelope towards the centrosomes. Turgay and colleagues found that the SUN proteins help to clear membranes from chromatin during NEBD. SUN proteins reside in the inner nuclear membrane and are part of a complex that connects the nucleus to the cytoskeleton. As seen in the images above, simultaneous depletion of both SUN1 and SUN2 (bottom timelapse) delayed removal of the nuclear envelope (green; chromosomes in red), when compared to control (top).  

Turgay, Y., Champion, L., Balazs, C., Held, M., Toso, A., Gerlich, D., Meraldi, P., & Kutay, U. (2014). SUN proteins facilitate the removal of membranes from chromatin during nuclear envelope breakdown The Journal of Cell Biology, 204 (7), 1099-1109 DOI: 10.1083/jcb.201310116

April 9, 2013

Our genome is chock full of so many things that aren’t even genes.  In fact, only about 2% of the human genome actually encodes protein sequences...mind blown, right?!  There are many different kinds of elements and domains within our genome that regulate gene expression through their roles in chromosome architecture and organization.  Today’s image is from a paper that describes the dynamics of one type of domain—the lamina associated domain.

The nuclear lamina is a protein layer that coats the inside nuclear membrane, and serves to anchor chromosomes.  Regions in the genome called lamina associated domains (LADs) specifically associate with the nuclear lamina.  LADs cover about 35-40% of the genome, suggesting that they may affect chromosome position and architecture.  A recent paper tracks LAD-nuclear lamina interactions throughout the cell cycle in single cells.  Kind and colleagues show that about 30% of LADs are positioned at the nuclear periphery.  LADs are stochastically positioned after mitosis, meaning that their position is not directly inherited.  In addition, these contacts are linked with gene expression and histone modifications.  In the images above, the nuclear lamina (blue) and mitotic spindle (red) are shown throughout the different stages of mitosis.  LADs (green) are rounded and at the nuclear periphery during prophase, and then become banded along chromosomes once the nuclear envelope breaks down (prometaphase and metaphase).  During cytokinesis, LADs are seen within the nucleus, but not yet positioned at the periphery.

Kind, J., Pagie, L., Ortabozkoyun, H., Boyle, S., de Vries, S., Janssen, H., Amendola, M., Nolen, L., Bickmore, W., & van Steensel, B. (2013). Single-Cell Dynamics of Genome-Nuclear Lamina Interactions Cell, 153 (1), 178-192 DOI: 10.1016/j.cell.2013.02.028
Copyright ©2013 Elsevier Ltd. All rights reserved.

March 7, 2013

DNA is not just shoved into the nucleus of a cell like a college boy’s clothes jammed into his closet (maybe that was just my husband?).  The organization of the nucleus has been getting more attention lately, and the results are pretty fascinating.  Today’s image is from a recent paper showing the association of a promoter to nuclear pores.

The organization of the nucleus may depend on the tethering of chromatin, or packaged DNA, to the nuclear envelope.  While some past results have suggested that actively expressed regions of chromatin are associated with the nuclear envelope in some organisms, other results have shown localization of actively expressed genes at internal nuclear speckles in other organisms.  A recent paper shows a link between the nuclear pore and a promoter, which is a region of DNA that initiates the expression of a nearby gene.  Rohner and colleagues found that the heat shock promoter hsp-16.2 interacts with the nuclear pore after heat shock, a type of stress, in worms.  Without heat shock, the promoter still maintained a perinuclear localization.  Using super-resolution structured illumination microscopy (SR-SIM) to achieve 100-nm resolution, Rohner and colleagues found that after heat shock, the promoter’s localization to the nuclear pore complex increased.  These results suggest that this stress-activated promoter may direct chromatin to the nuclear pores, where genes can be more easily accessed by transcription machinery.  In the image above, a 200-cell stage worm embryo is stained to show the localization of the hsp-16.2 promoter (green) at the nuclear envelope (red, DNA is in blue) under normal circumstances.

BONUS!!  Click here for a video of a 3D reconstruction of super-resolution images showing nuclear pores (green) and nuclear envelope (lamina, red).  

Rohner, S., Kalck, V., Wang, X., Ikegami, K., Lieb, J., Gasser, S., & Meister, P. (2013). Promoter- and RNA polymerase II-dependent hsp-16 gene association with nuclear pores in Caenorhabditis elegans originally published in the Journal of Cell Biology, 200 (5), 589-604 DOI: 10.1083/jcb.201207024

September 24, 2012

If you are a mitotic spindle aficionado or superfan, then you understand how the mitotic spindle has a life of its own. Information on the spindle’s composition, dynamics, and list of duties can fill entire volumes…and there is still so much we are learning about the spindle. Today’s image is from a paper that adds another chapter in our metaphorical mitotic spindle book.

The mitotic spindle is composed of dynamic microtubules and countless proteins that ensure chromosomes are equally segregated into identical daughter cells at the end of cell division. Because much of the structure of the spindle comes from the cytoplasm, the contributions from the nucleus have remained unclear. A recent paper identifies the long-elusive presence of a nuclear matrix—a viscous matrix of proteins that supports the formation and function of the spindle. The nuclear matrix is composed of nuclear proteins that reassemble after nuclear envelope breakdown into a pole-to-pole structure. By looking at the cell divisions in fruit flies, Yao and colleagues found that the nuclear matrix is distinct from spindle microtubules, and is found closely around chromosomes following drug treatments that depolymerized microtubules. The time-lapse images above show mitosis in the syncytial Drosophila embryo. The nuclear matrix (green, Chromator) has a spindle-like morphology as the microtubule spindle (red, tubulin) itself forms. 

Yao C, Rath U, Maiato H, Sharp D, Girton J, Johansen KM, & Johansen J (2012). A nuclear-derived proteinaceous matrix embeds the microtubule spindle apparatus during mitosis. Molecular biology of the cell, 23 (18), 3532-41 PMID: 22855526

September 11, 2012

If I had to describe myself as a cell, I would want to say something about being a fired-up neuron, or a nurturing nurse cell, or a chameleon stem cell. Realistically, though, I’d be a muscle fiber—painfully organized and precise. When things aren’t so organized I’d just be a twitching little muscle cell until my closet was organized by sleeve length, my pantry in alphabetical order, my various to-do lists organized by priority. A recent paper describes how the nuclei and organelles within a muscle fiber are so regularly spaced (but sadly does not describe any suggestions for my own personal organization).

Striated muscle fibers have densely packed myofibrils, which are the structures that make muscle contractions happen. The muscle fiber is a large multinucleated cell, meaning that there are more than one nucleus. These nuclei, as well as other organelles, must be evenly distributed along the length of the fiber. KASH domain proteins play a role in positioning nuclei and other organelles in various cell types, and a recent paper found that two KASH proteins play this role in muscle cells in the fruit fly. Elhanany-Tamir and colleagues found that these two proteins, called Klar and MSP-300, cooperate to ensure the even spacing of nuclei within a muscle fiber. These two proteins link a nuclear ring of MSP-300 to astral microtubules. Without either of these KASH proteins, the nuclear spacing is irregular. MSP-300 also is important in the spacing of other organelles—mitochondria and ER—within the muscle fiber. In the images above, muscle fibers from wild-type (left), msp-300 mutant, and klar mutant fruit fly larvae are stained for MSP-300 (red) and a nuclear marker (green). In the mutants, the MSP-300 nuclear ring (arrowheads) is dissociated from the nucleus.

Elhanany-Tamir H, Yu YV, Shnayder M, Jain A, Welte M, & Volk T (2012). Organelle positioning in muscles requires cooperation between two KASH proteins and microtubules. originally published in the Journal of Cell Biology, 198 (5), 833-46 PMID: 22927463

August 20, 2012

When you watch a cell going through mitosis, it looks like a smooth ballet performance—grace with impeccable timing and synchrony. In reality, there is a lot going on within the cell to make mitosis progress so smoothly, just like the ballet dancers’ actual physical exertion and concentration. Today’s image is from a paper that describes the timely removal of proteins from the nuclear envelope during mitosis.

The nuclear envelope (NE) is a double membrane that separates the cell’s genome from the rest of the cell, and permits transport of material into and out of the nucleus through multi-protein complexes called nuclear pores. During mitosis, the NE breaks down in order to allow chromosomes to attach to the mitotic spindle. Prior to NE breakdown, the nucleoporins that make up the nuclear pore complexes must be dissociated from the NE. A recent paper describes the timely removal of the nucleoporin NPP-3 near centrosomes at the onset of mitosis in early worm embryos. Hachet and colleagues found that centrosomes and the Aurora-A kinase AIR-1 are both required for removal of NPP-3 from the NE. In the images above, NPP-3 (right column, red in merged) is localized on the NE and is removed as mitosis progresses. NPP-3 removal begins near centrosomes (microtubules in green). 

Hachet V, Busso C, Toya M, Sugimoto A, Askjaer P, & Gönczy P (2012). The nucleoporin Nup205/NPP-3 is lost near centrosomes at mitotic onset and can modulate the timing of this process in Caenorhabditis elegans embryos. Molecular biology of the cell, 23 (16), 3111-21 PMID: 22740626

January 31, 2011

When many people think of membrane fusion, they often think of endocytosis and vesicle trafficking. Membrane fusion is a fascinating and dynamic process that is involved in so many processes in the cell, including nuclear envelope formation.

The nuclear envelope is composed of an inner and outer membrane that surrounds DNA and associated proteins. The nuclear envelope contains nuclear pores, which are channels that serve as sites of exchange between the nucleus and cytoplasm of the cell. The double membrane nature of the nucleus presents challenges in nuclear envelope and pore assembly that are overcome by membrane fusion events. A recent paper describes inner/outer membrane fusion during nuclear pore assembly and identifies an intermediate step in the process. By using cold temperatures that slow down nuclear pore formation in frog egg extracts, Fichtman and colleagues were able to clarify when and how fusion occurs between the inner and outer membranes. Specifically, the cold temperature treatment blocks inner/outer membrane fusion while allowing only the nuclear side of the pore to assemble. Images above show nuclear pore formation at room temperature (left set of images) and a cold temperature (right set of images). At room temperature, the nuclear membrane (green) and nuclear pores (red) form around chromatin (blue in merged images) after 15 minutes from the start of nuclear assembly. At cold temperatures, nuclear pores form more slowly and don’t appear on the nuclear envelope until 60 minutes.

Fichtman, B., Ramos, C., Rasala, B., Harel, A., & Forbes, D. (2010). Inner/Outer Nuclear Membrane Fusion in Nuclear Pore Assembly: Biochemical Demonstration and Molecular Analysis Molecular Biology of the Cell, 21 (23), 4197-4211 DOI: g/10.1091/mbc.E10-04-0309">10.1091/mbc.E10-04-0309