Three-dimensional organization of DNA

The three-dimensional organization of DNA in the nucleus influences our biology, from how our genome organized our cellular activity to how genes are passed from parents to children.

 X-rays and microscopy showed that the primary level of chromatin organization involves 147 bases of DNA spooling around proteins to form particles approximately 11 nanometers (nm) in diameter called nucleosomes.

These nucleosome "beads on a string" are then thought to fold into discrete fibers of increasing diameter (30, 120, 320 nm etc.), until they form chromosomes.

Researchers used ChromEMT to image and measure chromatin in resting human cells and during cell division (mitosis) when DNA is compacted into its most dense form -the 23 pairs of mitotic chromosomes that are the iconic image of the human genome.

Chromatin that has been extracted from the nucleus and subjected to processing in vitro - in test tubes - may not look like chromatin in an intact cell, so it is tremendously important to be able to see it in vivo.

Chromatin's packing density, and not some higher-order structure, that determines which areas of the genome are active and which are suppressed.

Controlling access to chromatin could be a useful approach to preventing, diagnosing and treating diseases like cancer.

Chromatin does not need to form discrete higher-order structures to fit in the nucleus, It's the packing density that could change and limit the accessibility of chromatin, providing a local and global structural basis through which different combinations of DNA sequences, nucleosome variations and modifications could be integrated in the nucleus to exquisitely fine-tune the functional activity and accessibility of our genomes.
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Gene editing kills HIV-1 infection in animals

A permanent cure for HIV infection remains difficult because the virus can hide in their reservoirs.

 Scientists at the Lewis Katz School of Medicine at Temple University (LKSOM) and the University of Pittsburgh show that they can remove HIV DNA from the genomes of living animals to prevent further infection.

They  demonstrate that HIV-1 replication can be removed completely and the virus eliminated from infected cells in animals with a powerful gene editing technology known as CRISPR/Cas9.

Previously, scientists used transgenic rat and mouse models with HIV-1 DNA incorporated into the genome of every tissue of the animals' bodies. They demonstrated that their strategy could delete the targeted fragments of HIV-1 from the genome in most tissues in the experimental animals.

In the new study, the team genetically inactivated HIV-1 in transgenic mice, reducing the RNA expression of viral genes by roughly 60 to 95 percent, confirming their earlier findings. They then tested their system in mice acutely infected with EcoHIV, the mouse equivalent of human HIV-1.

"During acute infection, HIV actively replicates. With EcoHIV mice, scientists were able to investigate the ability of the CRISPR/Cas9 strategy to block viral replication and potentially prevent systemic infection." The excision efficiency of their strategy reached 96 percent in EcoHIV mice, providing the first evidence for HIV-1 eradication by prophylactic treatment with a CRISPR/Cas9 system.

In the third animal model, latent HIV-1 infection was recapitulated in humanized mice engrafted with human immune cells, including T cells, followed by HIV-1 infection.

 "These animals carry latent HIV in the genomes of human T cells, where the virus can escape detection," Dr. Hu explained. Following a single treatment with CRISPR/Cas9, viral fragments were successfully excised from latently infected human cells embedded in mouse tissues and organs.

In all three animal models, the researchers utilized a recombinant adeno-associated viral (rAAV) vector delivery system based on a subtype known as AAV-DJ/8. "The AAV-DJ/8 subtype combines multiple serotypes, giving us a broader range of cell targets for the delivery of our CRISPR/Cas9 system,"