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Building a genomic GPS

UMMS scientists develop multicolored CRISPR/Cas9 labeling system to precisely measure the distance between chromosomes and genetic elements

2015-03-12
(Press-News.org) WORCESTER, MA - A new "app" for finding and mapping chromosomal loci using multicolored versions of CRISPR/Cas9, one of the hottest tools in biomedical research today, has been developed by scientists at the University of Massachusetts Medical School. This labeling system, details of which were published in PNAS and first presented at the American Society for Cell Biology/International Federation for Cell Biology annual meeting in Philadelphia in December, could be a key to understanding the spatial and temporal regulation of gene expression by allowing researchers to measure the precise linear distance between two known points on different chromosomes or two locations on the same chromosome in live human cells.

The nucleus of every cell in our bodies (with the exception of gametes and red blood cells) must pack into it 23 pairs of chromosomes, tight bundles of extremely long strands of DNA wound around protein knobs. For a gene to be transcribed and expressed, it must be accessible on the chromosome. Scientists have long suspected that the position of a chromosome within the nucleus affects gene accessibility and plays a critical role in everything from embryonic development to cancer.

Knowing the location and the intra-nuclear conformation of chromosomes is critical to understanding how genes actually work because the human cell nucleus is a very crowded place, according to study authors Thoru Pederson, PhD, professor of biochemistry and molecular pharmacology, and research specialist Hanhui Ma, PhD, at UMass Medical School.

By deploying pairs of fluorescent tags from their three-color system, Pederson and colleagues showed that it's possible to plot where a chromosome is inside the cell nucleus and where it is in relation to other chromosomes. Their CRISPR app can also measure the distance between two points on the same chromosome, giving a read-out of chromosome compaction, which is a key factor in gene expression.

Precisely locating chromosomes in the nucleus of living cells has been a holy grail in cell biology since 1968, when Joseph Gall and Mary Lou Pardue first demonstrated the detection of specific loci. That discovery helped open the era of genetic testing, but the early techniques required "fixed," i.e. dead, cells. In the intervening years, researchers have adapted new methods to probe live cells including transcription activator-like effectors (TALEs), which Ma and Pederson had recently introduced for lighting up genetic loci in living cells. But they subsequently came to believe that the rapidly emerging CRISPR system promised a more accurate map of a living nucleus and would be easier for scientists to employ.

Using their multicolored system, they were able to determine common locations for several chromosomes. Among their findings was that gene-rich chromosome 19 tends to be located in the middle of the nucleus, whereas gene-poor chromosome 18 is at the periphery. Also staying close to the center of the nucleus were chromosome 17 and five of the so-called acrocentric chromosomes, which have their distinctive centromeres close to the ends of one arm. One of the five acrocentrics is chromosome 21; the presence of an extra copy of chromosome 21 is called trisomy 21, and is the diagnostic marker for Down syndrome. Meanwhile, chromosomes 3 and 7 were at or close to the nuclear periphery.

The researchers were also able to distinguish the different locations for each diploid copy of the genes involved in organizing the nucleolus. The researchers say that they have plans to further tweak their two-color technique to study translocations--the abnormal switching of chromosomal segments--in human tumor cells.

Ultimately, the GPS app will provides scientists a new toolkit for "studying the 4D nucleome and the regulation of eukaryotic gene expression across a broad landscape of cell types and stages of development, differentiation and human disease," the authors state in their publication.

INFORMATION:

The published study was funded in part by the U.S. National Science Foundation.

About the University of Massachusetts Medical School The University of Massachusetts Medical School (UMMS), one of five campuses of the University system, comprises the School of Medicine, the Graduate School of Biomedical Sciences, the Graduate School of Nursing, a thriving research enterprise and an innovative public service initiative, Commonwealth Medicine. Its mission is to advance the health of the people of the commonwealth through pioneering education, research, public service and health care delivery with its clinical partner, UMass Memorial Health Care. In doing so, it has built a reputation as a world-class research institution and as a leader in primary care education. The Medical School attracts more than $240 million annually in research funding, placing it among the top 50 medical schools in the nation. In 2006, UMMS's Craig C. Mello, PhD, Howard Hughes Medical Institute Investigator and the Blais University Chair in Molecular Medicine, was awarded the Nobel Prize in Physiology or Medicine, along with colleague Andrew Z. Fire, PhD, of Stanford University, for their discoveries related to RNA interference (RNAi). The 2013 opening of the Albert Sherman Center ushered in a new era of biomedical research and education on campus. Designed to maximize collaboration across fields, the Sherman Center is home to scientists pursuing novel research in emerging scientific fields with the goal of translating new discoveries into innovative therapies for human diseases.



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[Press-News.org] Building a genomic GPS
UMMS scientists develop multicolored CRISPR/Cas9 labeling system to precisely measure the distance between chromosomes and genetic elements