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 19, 2013

After a lovely Spring Break with my family, HighMag is back in action.  As an oldie, Spring Break takes on a whole new meaning than it did years ago…this pregnant lady didn’t have one single beer!

 Cancer is a series of cellular mistakes, mistakes that are too far gone to fix without the help of the medical field.  Understanding the mistakes at the most basic cellular level is key to fighting the war on cancer, and a recent paper is a fine example of this.

A great model for understanding cell polarity is the intestinal epithelial sheet of cells that provides a barrier between the inside of the intestine and the body.  The different polarized domains – apical and basal – each have a discrete set of adhesion and membrane proteins trafficked to them.  One membrane trafficking protein, Rab25, is a tumor suppressor for colon cancer in both humans and mice.  A recent paper investigates the link between Rab25 and the polarized intestinal cells involved in colon cancer.  Krishnan and colleagues found that a reduction of Rab25 levels in cultured endothelial cells resulted in increased cell invasion and a loss of certain integrins, adhesion proteins, at the plasma membrane.  Rab25 loss also affected the transcription of several genes including the transcription factor ETV4, suggesting that Rab25’s effect on cell polarity is through gene regulation.  The scanning electron images above show brush border microvilli, the fingerlike-projections seen in intestinal epithelial sheets that serve to increase absorption of nutrients from the intestine.  In Rab25-reduced cells (middle row), the brush border is sparsely-packed and disorganized when compared to control cells (top row).  Bottom row shows Rab25-reduced cells in which rabbit Rab25 was reintroduced, and the rescued brush border is more organized.

Krishnan M, Lapierre LA, Knowles BC, & Goldenring JR (2013). Rab25 regulates integrin expression in polarized colonic epithelial cells. Molecular biology of the cell, 24 (6), 818-31 PMID: 23345591

February 15, 2013

The regulation of genes occurs at many levels, one of the first of which is physical access to a gene.  The way DNA is packaged may or may not allow transcription machinery from even getting to an area of the genome.  Understanding these modifications in each cell type and location is important, but difficult within a tissue or tumor of different cell types.  A recent paper tackles this problem with a new method.

Histones are proteins that package DNA to condense its size and to aid in gene regulation.  Histones can be modified in many different ways, and these modifications affect how accessible or inaccessible a certain region of the genome is to gene transcription.  Chromatin immunoprecipitation, the current technique to understand histone modifications at different gene sites, does not provide information on histone modifications of single cells within a complex environment of different cell types.  A recent paper describes a new technique that allows the visualization of histone modifications at single-cell resolution within a fixed tissue.  Using this technique, Gomez and colleagues tracked one specific histone modification, dimethylation of lysine 4 of histone H3 (H3K4me2) at the genetic loci for MYH11 in smooth muscle cells (SMC).  SMC-containing tissues contain other cell types, and even non-SMC tissues contain some SMCs due to vascularization.  In the images above, MYH11 H3K4me2 modifications (red, arrows) were found only in SMCs (green cells) in sections of human carotid artery tissue (DNA in blue).  Higher magnification images are bottom row.

Gomez, D., Shankman, L., Nguyen, A., & Owens, G. (2013). Detection of histone modifications at specific gene loci in single cells in histological sections Nature Methods, 10 (2), 171-177 DOI: 10.1038/nmeth.2332
Adapted by permission from Macmillan Publishers Ltd, copyright ©2013 

December 13, 2012

DNA is decorated more beautifully than a Christmas tree, but the decorations are not just for looks—they serve the important function of regulating which genes get expressed, and when. Today’s image is from a paper describing the role of a protein that affects gene expression by regulating the accessibility of some regions of the DNA.

Chromatin is the packaged DNA and associated proteins found in the nucleus. Certain regions of chromatin are packaged or modified to make the underlying genes more or less accessible to the transcription machinery that results in gene expression. A recent paper describes the identification of UpSET, a fruit fly protein that binds to promoter regions of transcriptionally active genes. The upSET gene resembles the mammalian gene MLL5, which is found in a region frequently deleted in a subset of leukemias. In the absence of UpSET, according to Rincon-Arano and colleagues, cells have increased chromatin accessibility and express genes that normally flank the regions of UpSET binding. In the images above, fruit fly polytene chromosomes (blue) show staining for UpSET (green) in gene-rich regions.

Rincon-Arano, H., Halow, J., Delrow, J., Parkhurst, S., & Groudine, M. (2012). UpSET Recruits HDAC Complexes and Restricts Chromatin Accessibility and Acetylation at Promoter Regions Cell, 151 (6), 1214-1228 DOI: 10.1016/j.cell.2012.11.009 
Copyright ©2012 Elsevier Ltd. All rights reserved.

May 21, 2012

Whenever we go on a trip, my long-suffering husband quietly puts our luggage next the car and slinks away, trembling and twitching.  He knows a mad-woman is ready to pack the trunk, playing luggage-Tetris until it all fits and speaking in tongues.  Seriously, though, I’m freaking awesome.  That said, I don’t envy the insane packing that a cell must accomplish to jam all of that DNA into neat little chromosomes ready for their own cell division road trip.  A recent paper helps us understand how that happens at the centromere.

Centromeres are the regions on chromosomes that bind sister chromatids together and serve as the sites of kinetochore assembly during mitosis.  The presence of the protein CENP-A is a hallmark of centromere location, as it is a histone H3 variant that helps package and compact centromeric DNA.  It was previously presumed that CENP-A was passed down to daughter cells epigenetically, inherited from previous cell divisions, but a recent paper shows that this is not the case in the nematode worm C. elegans.  According to Gassmann and colleagues, pre-existing CENP-A is not required for CENP-A localization to centromeres in subsequent divisions.  In fact, CENP-A is unloaded from centromeres at one point in oogenesis, the production of eggs, and later reloaded onto centromeres.  By mapping the location of CENP-A in the genome, Gassmann and colleagues found that regions of transcribed genes are regions where CENP-A is excluded, a pattern that changes when germline gene transcription switches to embryonic gene transcription.  In the images above, the C. elegans germline is labeled to show chromosomes (top image) and the location of CENP-A (bottom).  CENP-A is lost from chromosomes during the pachytene stage of meiosis and later reloaded onto chromosomes during diplotene, and is not found in sperm.  

Gassmann, R., Rechtsteiner, A., Yuen, K., Muroyama, A., Egelhofer, T., Gaydos, L., Barron, F., Maddox, P., Essex, A., Monen, J., Ercan, S., Lieb, J., Oegema, K., Strome, S., & Desai, A. (2012). An inverse relationship to germline transcription defines centromeric chromatin in C. elegans Nature, 484 (7395), 534-537 DOI: 10.1038/nature10973
Adapted by permission from Macmillan Publishers Ltd, copyright ©2012

January 27, 2011

Just when you think you understand how a gene works…BAM! Alternative splicing shows up and reminds us that there is so much yet to learn, even about a gene as well-studied as formin.

Genes get transcribed into RNA, which gets translated into proteins. After transcription of a gene, different regions of RNA called exons and introns are either connected together (exons) or removed (introns) in a process called splicing. Some genes undergo a process called “alternative splicing” that allows one single gene the ability to splice, or connect, the exons in multiple ways that result in different protein isoforms. A recent paper describes a previously-unidentified isoform of FHOD3 formin, an actin-nucleating protein found at high levels in the heart. This new isoform includes an alternative exon in certain muscle tissues, and this exon contains a phosphorylation site that allows an additional level of regulation of FHOD3. Images above are of neonatal rat heart cells with or without this alternative exon. The presence of the alternative exon (bottom cell) directs formin (left images, green in merged) to myofibrils (middle images, red in merged), which are the repetitive and contractile structures in muscle cells. Without the alternative exon, formin is found in aggregates in the cytoplasm (top cell).

Iskratsch, T., Lange, S., Dwyer, J., Kho, A., Remedios, C., & Ehler, E. (2010). Formin follows function: a muscle-specific isoform of FHOD3 is regulated by CK2 phosphorylation and promotes myofibril maintenance originally published in The Journal of Cell Biology, 191 (6), 1159-1172 DOI: 10.1083/jcb.201005060