Dr. Sujan Mamidi
HudsonAlpha Institute for Biotechnology
The advent of whole-genome sequencing revolutionized our understanding of biological systems. The early 2000s marks the beginning of human genomics when the first human genome sequencing was published in 2003. The availability of advanced sequencing techniques coupled with lower cost for billions of bases completely changed the scale and our understanding of biology. Over the years in addition to whole-genome DNA sequencing, we had RNA sequencing (the study is termed Transcriptomics) to understand the gene function. For this messenger RNA is extracted from different tissues to understand the expression patterns in each of the tissues that vary a lot among them. This also led to the discovery of alternate splice sites where a gene can be expressed in different forms in different cells/tissues. Besides, multiple conditions led to varied expression patterns of the genes.
However, the study limited our understanding to only a few tissues and conditions, and to know whether a gene is expressed or not and if it does, the structure of isoform. But knowing wherein the tissue is the gene products and what cell types, help understand genes better which can have many applications. A new method was developed, where tissue is expanded, individual molecules of mRNA are labeled, and then sequenced. This approach provides an instant snapshot of the genes expressed in different parts of the cell, and we can learn more about how gene expression is influenced by the cell location and its interaction with neighboring cells. This technique termed expansion sequencing (ExSeq) is published in Science, along with its use in the mouse brain and human tumor samples.
This is an extension of the previous method devised in 2015 of expanding tissues and then imaging them. This expansion was achieved by embedding water-absorbent polymers into a tissue that expanded the tissue about 100 times or more and then obtaining high-resolution images of the tissues using a light microscope. Also, in 2014 an RNA sequencing technique known as FISSEQ (fluorescent in situ sequencing) was developed, that allowed thousands of mRNA molecules to be located and sequenced. Combining these 2 approaches, the new technique, expansion sequencing (ExSeq) was created. Once the tissue is expanded, RNA is labeled and sequenced. This allows us to pinpoint the molecules’ locations within cells. Expanding the tissue offers a higher-resolution look at the RNA in cells, and it makes it easier to sequence those RNA molecules independently. It is estimated that in a given sample, about 20 and 50 percent of all of the genes present in tissue can be sequenced.
In the mouse hippocampus, the researchers discovered mRNA containing introns and mRNA encoding transcription factors in dendrites. Also, they were able to classify neurons based on 42 genes expressed. This study also revealed that cell types behave differently in different locations within a tumor. For example, the B cells near tumor cells expressed certain inflammatory genes at a higher level than those farther from tumor cells.
This is a significant step for our efforts to treat complex diseases, such as Alzheimer’s and cancer. Using this new technology, we will be able to identify genes and also their precise location just like coordinates on a geographical map.