Unveiling the role of Daam1 gene in intestinal stem cell fate decisions

Stem cells can differentiate to interchange dead and damaged cells. But how do stem cells resolve which form of cell to turn out to be in a given situation? Using intestinal organoids, the group of Bon-Kyoung Koo at IMBA and the Institute for Basic Science identified a brand new gene, Daam1, that plays a vital role, switching on the event of secretory cells within the intestine. This finding, published on November 24 in Science Advances, opens recent perspectives in cancer research.

Our bodies are, in some ways, like cars – to maintain functioning, they have to be checked and repaired commonly. Within the case of our bodies, any cells which are damaged or dead have to be replaced to maintain organs functioning. This alternative occurs due to tissue-resident adult stem cells. In contrast with embryonic stem cells, which may form any cell type within the body, adult stem cells will only form the cell types which are present in the tissue they belong to. But how do tissue-specific stem cells know which cell type to offer rise to? Gabriele Colozza, a postdoctoral researcher within the lab of Bon-Kyoung Koo at IMBA – now director on the Center for Genome Engineering, Institute for Basic Science in South Korea – decided to research this query using intestinal stem cells.

Intestines – a relentless construction site

“In our intestines, cells are exposed to extreme conditions”, Colozza explains. Mechanical wear and tear, but additionally digestive enzymes and ranging pH values all affect intestinal cells. In turn, stem cells within the intestine’s mucosa differentiate to form recent intestinal cells. “Damaged cells have to get replaced, but it surely is a fragile balance between stem cell renewal and differentiation into other cell types: uncontrolled stem cell proliferation may result in tumor formation; however, if too many stem cells differentiate, the tissue can be depleted of stem cells and ultimately unable to self-renew.”

This balance is delicately tuned by signaling pathways and feedback loops, which permit cells to speak with one another. One essential pathway is known as Wnt. The Wnt pathway is thought for its role in embryonic development, and if left unchecked, an overactive Wnt pathway can result in excessive cell division and the formation of tumors.

Molecular partner identified

A well known antagonist of Wnt signaling – keeping Wnt in check – is Rnf43, which was originally identified by Bon-Kyoung Koo. Prior to this study, Rnf43 was known to focus on the Wnt receptor Frizzled and mark it for degradation. “We desired to understand how Rnf43 works, and in addition what – in turn – controls Rnf43 and helps it to manage Wnt signaling.” From earlier research, the scientists knew that Rnf43 by itself was not sufficient to interrupt down the Wnt receptor Frizzled, which sits within the plasma membrane. “In our project, we used biochemical assays to discover which proteins interact with Rnf43.” A key partner of Rnf43 turned out to be the protein Daam1.

To grasp how Daam1 regulates Rnf43 and affects the tissues it acts in, Colozza turned to intestinal organoids. “We found that Daam1 is required for Rnf43 to be energetic, so for Rnf43 to manage Wnt signaling in any respect. Further work in cells showed Rnf43 needs Daam1 to maneuver the Wnt receptor Frizzled into vesicles called endosomes. From the endosomes, Frizzled is shuttled to the lysosomes where it’s degraded, dampening Wnt signaling”, Colozza adds.

Intestinal organoids are three-dimensional cell cultures grown from adult intestinal stem cells, allowing the researchers to mimic the intestinal mucosa. For Colozza, organoids were a chance to know how Rnf43 and Daam1 affect the fragile balance of stem cell renewal and differentiation within the intestine. “We found that once we knock-out Rnf43 or Daam1, the organoids grow into tumor-like structures. These tumor-like organoids carry on growing, even when we withdraw the expansion aspects they sometimes depend upon, comparable to R-spondin.”

Switching on Paneth cell formation

When Colozza followed up this lead to mouse tissue, the researchers were in for a surprise. “When Rnf43 was missing, the intestines grew tumors – as expected. But when Daam1 was missing, no tumors grew. We were puzzled by this striking difference: how can the loss of things in the identical pathway, that behave similarly in organoids, result in such different outcomes?”

Looking closely on the intestines, Colozza saw that intestines lacking Rnf43 were stuffed with a particular form of secretory cells, the Paneth cells. Intestines lacking Daam1, however, contained no extra Paneth cells. Paneth cells secrete growth aspects, comparable to Wnt, that stimulate cell division. “Daam1 is required for the efficient formation of Paneth cells. When Daam1 is energetic, stem cells differentiate to form Paneth cells. When Daam1 is just not energetic, the stem cells differentiate into one other cell type.”

Tumors modify their area of interest to grow

This link between the molecular results and Paneth cells explains the puzzling difference between intestines and organoids. “In organoid culture, we scientists provide growth aspects, so the knockout of each Rnf43 and Daam1 result in tumor-like organoids. But within the intestine, there isn’t any little scientist providing growth aspects. As an alternative, Paneth cells provide growth aspects, like Wnt, and create the proper conditions for stem cells to survive and divide. When Paneth cells are lacking – comparable to when Daam1 is just not energetic to drive cells into becoming Paneth cells – stem cells is not going to divide much. But when there are too many Paneth cells – comparable to in intestines lacking Rnf43 – the excessive growth aspects can contribute to the formation of tumors.”

Colozza’s and colleagues’ study is the primary genetic proof that Daam1, a member of the non-canonical Wnt pathway, is vital for specifying Paneth cells, and directly involved in the event of this significant secretory cell. The outcomes also make clear the importance of the stem cell area of interest. “We show that tumor cells modify their microenvironment, and influence their supporting environment in order that they’ll grow higher.”