Epigenetics and ASXL-related disorders

Introduction

ASXL-related disorders, including Bohring-Opitz Syndrome (ASXL1), Shashi-Pena Syndrome (ASXL2), and Bainbridge-Ropers Syndrome (ASXL3), are epigenetic disorders. The ASXL genes give instructions for proteins that play a significant role in regulating how our 30 billion base genetic code “DNA” is packaged within each cell in our body.  This article describes DNA structure and packaging, provides a detailed explanation of epigenetics, and the role of ASXL genes in epigenetics.

Editorial note for families: Epigenetics is really complicated (even for scientists)! We recommend watching some of the videos linked below and referencing the illustrations to help your understanding, and you may need to read this article several times.

The structure of DNA

DNA: base pairs in a double-helix structure

Before we can dive into epigenetics, there are certain “nitty-gritty” aspects of the structure of DNA that are important to understand. DNA, the genetic code that provides instructions for our bodies, is made up of four paired bases. The bases are called adenine, thymine, cytosine and guanine.  (You may see these bases abbreviated to the first letter of their names, such as “A, T, C, G.”)

The bonds between these base pairs create the double-helix structure of DNA. A double helix looks like a twisted ladder, with each rung being a base pair bond. The sequence of the base pairs moving down the ladder is the genetic code. This genetic code is read and translated to eventually make proteins that are essential to a functioning human body. Gene expression is when the information stored in our DNA is translated into proteins and other molecules, ultimately, leading to our physical characteristics (also referred to as phenotype).

Compaction: How a long strand of DNA to fits in a cell

The challenge with the structure of DNA is that the double-helix is approximately 30 billion base pairs long—too long to fit into a single cell as a long strand. In fact, if you were to uncoil and stretch the DNA in a cell, it would reach six feet long. So, how does each individual cell in your body contain six feet of DNA? The answer lies in several organized levels of compaction.

In the first level of compaction, the negatively charged DNA double helix continuously wraps around a group of eight positively charged proteins called histones, like a spool of thread. This single spool is known as a nucleosome.  

In the second level, these nucleosomes wind up close together and the resulting tightly bound package of histone spools is called chromatin.  

In the third, fourth and fifth level, chromatin winds itself up, then winds itself again and again so tightly that it folds onto itself to create neatly packed chromosomes (see picture). Humans have 22 pairs of numbered chromosomes (autosomes) and a pair of sex chromosomes (XX or XY) for a total of 46 chromosomes.  These are the X and Y shaped chromosomes that we all learned about in high school biology, the ones you inherit from your parents.

Epigenetics 101

Overview

Epigenetics literally means “above the genes” (in Latin, “epi” means “above”). In epigenetics, a change can happen in the expression of a gene without disrupting the underlying genetic sequence. Epigenetic mechanisms control chromatin (the tightly wound structures of DNA) by giving it instructions in the form of chemical tags or marks that tell the chromatin structure when to open or close so the DNA can be read and used to make the right proteins at the right time and place (cell in a specific organ or tissue).

The ASXL genes are part of this chromatin regulatory system that controls the epigenetic processes that open and close chromatin for the DNA to be accessed and read.

Epigenetic regulation: Opening and closing chromatin

As you recall from earlier in this article, DNA carries the instructions to make the proteins necessary for our bodies to function. When chromatin is open, or loosely packed, the DNA can be read and translated into instructions used to make the necessary proteins. (See the illustration). When chromatin is closed, or tightly packed together, the instructions can’t be accessed to be read or followed.  It’s important to remember that whether or not the DNA book of instructions is open or closed does not change the DNA itself – it just changes if the DNA can be accessed.

It may help to think of DNA as a textbook of instructions. Your teacher may tell you that for today’s assignment you only need to read pages 5-15 and 30-60 in your textbook. Imagine your teacher helps you by paperclipping together the pages that you don’t need to read. All the pages in the book are all still there, but now you can only read the pages necessary for this lesson. The epigenetic regulators are like the teacher who paperclips certain pages together. Those pages of instructions are now “closed”, and the other pages are “open.” This ensures that you read the right pages for today’s lesson.

Our DNA also carries the code for the proteins that function in and control epigenetic processes. The ASXL genes carry the code for ASXL proteins, which are epigenetic regulators, in that they impact the way that other genes are expressed through placing tags signaling the opening or closing of chromatin. When there is a mutation in an ASXL gene and the protein is not made or is made incorrectly, other genes will be expressed differently because the ASXL protein is not present for its role in controlling when and how other parts of the genetic code can be accessed and read. 

Tags and tails: DNA methylation and histone modification control the opening and closing of chromatin

The processes the body uses to regulate and organize the compaction (the winding and folding of DNA) and therefore whether the DNA is “open” or “closed” are at the core of epigenetics. The two main processes to give the “open” and “close” instructions are called DNA methylation and histone modification. These methods involve the placement or removal of chemical tags that determine whether conditions are right for reading the DNA and forming functional proteins based on the DNA instructions. This is known as gene expression.  

Again, these tags do not change the DNA itself; rather, they change whether the DNA instructions are read (and therefore whether the proteins turn out or not) by “paperclipping” the DNA instructions together in places. The collection of tags (or paperclips) all throughout our genetic code are referred to as the epigenome.

In the first type of tag, DNA methylation, a methyl group (one carbon and three hydrogen atoms) is added directly to the DNA sequence at specific sites. When a collection of these tags are placed on the DNA it signals whether the gene should be read or ignored, effectively turn the gene “on” or “off.” Researchers look to identify patterns in where these methyl tags are placed on the DNA to better understand how the tags work and how their placement affects a person's symptoms and characteristics. Unique patterns in how the tags are placed are known as DNA methylation signatures which can be specific to rare genetic disorders, such as ASXL-related disorders. (We know through research that the ARRE Foundation helped fund that Bohring-Opitz Syndrome has a distinct DNA methylation signature and additional studies are underway to determine if this is also true for Bainbridge-Ropers Syndrome and Shashi-Pena Syndrome.)

The other most common type of tag is the addition of different chemicals to histone protein “tails.” Histone proteins (the spools around which DNA wraps) have tails which are made up of a small chain of amino acids (the blocks that make protein). These tails aren’t just cute appendages; rather, they are critical to how and when chromatin is compacted.  

Depending on factors like where the tag is on the tail and which chemical the tag is made of, they either push the nucleosomes apart so that the DNA can be read or pull them tighter together and, like the paperclip from before, prevent them from being read.  This is called histone modification. Some of the chemical tags that can attach to these tails are methyl groups, acetyl groups, ubiquitin, and phosphoryl. Scientists often study each epigenetic mark and its effects separately, for example acetyl groups cause the DNA to be “open.” Studying and understanding the combined effect of epigenetic marks is more complex and challenging. Many scientists are currently studying epigenetics changes to learn more.   

Whether DNA methylation or histone modification are used, both methods leave tags and both methods impact whether the DNA is “open or “closed.” This means that either one can affect what proteins are formed and DNA “instructions” are followed.

PR-DUB complex: What we know about epigenetic complexes and ASXL-related disorders

There are three main epigenetic complexes that add and remove these chemical tags directly from the DNA (DNA methylation) and from the histone tails (histone modification). These complexes are made up of multiple proteins put together forming an assembly of proteins. They are called: PR-DUB (said “P” “R” “dub”), PRC1 and PRC2. These complexes are like the teacher in our above example of the textbook. The complexes are responsible for putting the paperclips on and taking them off the pages as needed so the book can open and close to the correct pages.

The functional ASXL1, ASXL2, and ASXL3 proteins are all part of the PR-DUB complex. These proteins combine with BAP1 and other proteins to play a crucial role in both fetal development and the ongoing regulation of our genes. In individuals who have ASXL-related disorders, the functions of the PR-DUB complex are affected due to the mutations/changes in the ASXL genes that cause the ASXL proteins not to be produced or a produced with abnormalities to structure and function.

The PR-DUB complex regulates a specific tag called H2AK119ub. The naming indicates the tag is placed on histone 2A at the amino acid lysine position 119, and the mark is a ubiquitin mark indicated by the (ub). Due to mutations in ASXL genes, the PR-DUB complex may not function correctly which leads to the histone modifications (tags on the histone tails) not being placed correctly. As a result, the cell can no longer access the correct DNA instructions at the right time and the cell is unable to function normally. Although these changes happen at a cellular level, they are partly responsible for the multi-organ symptoms of ASXL-related disorders.

Researchers are working to better understand exactly what happens in ASXL-related disorders. The hope is that if we can gain further insight into these chemical and epigenetic changes, more effective therapies can be developed that target the specific chromatin modification mechanisms of the ASXL genes.  

Suggestions for additional learning

Acknowledgements

Thank you to Elian Silverman, a genetic counseling student, who wrote this article. We are are grateful to Debbie Requesens and the JumpStart Program of the Orphan Disease Center for pairing Elian with us for this project.

We also extend our deep gratitude to Zain Awamleh, PhD for reviewing this article.