Newly Identified Enzyme Enables Lifelong Sperm Production

The enzyme DOT1L, a stem cell self-renewal factor, has been found to be essential for mice to continue generating sperm throughout maturity, according to research from the University of Pennsylvania.

Contrary to women who are born with all of their eggs, men may continue to produce sperm throughout their adult lives. They must continuously replenish the spermatogonial stem cells in order to produce sperm.

This stem cell renewal is dependent on a recently discovered stem cell self-renewal component known as DOT1L, according to research by Jeremy Wang of the University of Pennsylvania School of Veterinary Medicine and associates. The researchers showed that DOT1L-deficient animals are unable to maintain spermatogonial stem cells, which impairs their capacity to continuously produce sperm.

The discovery, which was published in the journal Genes and Development, brings the number of stem cell renewal factors that have already been found by researchers up to a few more.

The fact that mice lacking DOT1L were unable to continue producing sperm allowed researchers to pinpoint this novel component, says Wang, the Ralph L. Brinster President's Distinguished Professor at Penn Vet and a corresponding author on the study.

In addition to assisting in our understanding of the biology of adult germline stem cells, the discovery of this crucial component may one day enable us to rewire somatic cells, such as the fibroblasts of the skin, to differentiate into germline stem cells, effectively producing a gamete in a petri dish. The future of fertility treatment lies there.                                                                                              

Spermatogonial stem cells exhaust themselves when the enzyme DOT1L isn't working, which prevents sperm cell growth. A Penn Vet team discovered that DOT1L plays a vital role in stem cell self-renewal, placing it in exclusive company as one of just a select few recognized variables. Thanks to Jeremy Wang

The researchers unintentionally found that DOT1L has a role in stem cell self-renewal. Mice having a mutant variant of DOT1L in every cell do not survive through the embryonic stage despite the gene being extensively expressed. However, DOT1L's genetic expression patterns led Wang and associates to postulate that it might be involved in meiosis, the process of cell division that yields sperm and eggs. In order to find out what would happen if they altered the gene exclusively in these germ cells, they decided to do some research.

The animals appeared to be healthy and lived when we performed this, according to Wang. When we paid closer attention, however, we discovered that although the mice with the mutant DOT1L in their germ cells were able to produce sperm for the first time, after that the stem cells ran out and the mice lost all of their germ cells.

There could be other issues causing this decline in sperm production. However, a number of lines of research suggested a connection between DOT1L and a lack of stem cell self-renewal. The scientists discovered that the mice sequentially lost the ability to produce spermatogonia, spermatocytes, round spermatids, and elongated spermatids, as well as other sperm formation stages.

In a second experiment, the researchers looked at what would happen if DOT1L was inactivated in germ cells throughout adulthood rather than at birth. Wang and colleagues found that the identical progressive loss of sperm development they had seen in the mice born without DOT1L in their germ cells occurred as soon as the DOT1L loss was activated.

DOT1L has previously been investigated in relation to leukemia by other scientific teams. Malignancy may result from the gene's overexpression in the blood cell progenitors. It was discovered through that line of research that DOT1L functions as a histone methyltransferase, an enzyme that modifies the expression of genes by adding a methyl group to histones.

Wang and his team treated spermatogonial stem cells with a substance that inhibits the methyltransferase activity of DOT1L to test if the same mechanism was in charge of the outcomes they had seen in sperm formation. The spermatogonia-producing capacity of the stem cells was greatly diminished as a result. Additionally, the therapy reduced stem cells' capacity to methylate histones. Additionally, the activity of the animals' spermatogonial stem cells was reduced in half after these treated stem cells were implanted into otherwise healthy mice.

The scientists discovered that DOT1L appeared to be controlling the Hoxc gene family, transcription factors that are important in controlling the expression of a variety of other genes.

We believe that DOT1L methylates these Hoxc genes to encourage their expression, according to Wang. "These transcription factors most likely aid in the process of stem cell regeneration. Future directions for our work include learning more about that.

Longer term objectives include using DOT1L and other germline stem cell self-renewal factors to assist individuals with reproductive issues. The idea is to generate germ cells from scratch.

In vitro gametogenesis is the field's future, according to Wang. One of the steps is to reprogram somatic cells to become spermatogonial stem cells. The next step would be to figure out how to induce meiosis in those cells. Although we're still working out the details of how to complete this multi-phase process, recognizing this self-renewal component moves us one step closer.

The China Scholarship Council, the National Natural Science Foundation of China, the Japan Society for the Promotion of Science, and the Eunice Kennedy Shriver National Institute of Child Health and Human Development all provided funding for the study.

We advise using frozen testicular tissue, which can produce viable sperm and be reimplanted after 20 years.

By UNIVERSITY OF PENNSYLVANIA