THE DROSOPHILA TESTIS NICHE
The fly testis is an ideal model to study stem cell interactions as the niche and resident stem cells are easily identified and in stereotyped positions. The testis is a coiled tube with the niche located in the apical tip (left). The niche contains a group of somatic cells called the hub that anchor the two stem cell populations, the germline stem cells and somatic stem cells. After mitosis, one GSC daughter is retained in the niche and the other, the gonialblast, is displaced and begins to differentiate. This Gb absolutely MUST association with exactly two daughters of the somatic stem cell lineage, the cyst cells, or the Gb will fail to differentiate and sperm production will cease. We use this system to study stem cell coordination within the niche--a critical, but largely unexplored question in stem cell biology.
HOW DOES THE NICHE COORDINATE SOMATIC AND GERMLINE STEM CELLS?
A 2:1 RATIO OF SOMA:GERMLINE IS NECESSARY FOR GERM CELL DIFFERENTIATION AND SPERM PRODUCTION
Every one gonialblast produced by a GSC must be encysted by precisely two cyst cells generated by the somatic stem cells. As the cyst cells themselves do not divide, it means that this required ratio of two somatic cells for everyone one germ cell MUST be established in the niche.
STEM CELLS DO NOT COORDINATE DAUGHTER CELL PRODUCTION THROUGH SYNCHRONOUS DIVISIONS
The simplest model to explain coordinated production of daughter cells is synchronous divisions of the two stem cell populations (top). Every mitotic GSC would be flanked by two mitotic somatic stem cells--the end result would be production of 2:1 cyst cells:gonialblast. However, we have found that this synchrony DOES NOT EXIST in the testis niche. A GSC in mitosis (white nucleus) is NEVER flanked by two somatic stem cells in mitosis (bottom).
COMPLETION OF GSC CYTOKINESIS IS EXTREMELY DELAYED
Instead, we believe that a unique aspect of GSC cell biology--an extremely delayed completion of cytokinesis--serves to coordinate daughter cell RELEASE from the niche. Typical cells complete cytokinesis through the act of abscission, or severing of daughter cell membranes, at the end of mitosis or very early in G1. By contrast, GSCs do not undergo abscission until very late in the following cell cycle, sometime during G2.
MODIFIED STEM CELL CYTOKINESIS
The Lenhart lab has established a powerful live imaging system to visualize stem cells within the endogenous niche for over 20 hours--more than an entire GSC cell cycle. Using this approach and labeling actin in germ cells (green) and myosin in all cells (magenta) we have successfully observed the entire process of GSC cytokinesis from mitosis through abscission.
TWO PHASES OF GSC CYTOKINESIS
Live imaging reveals that GSCs progress through two phases of cytokinesis. After contractile ring disassembly in mitosis, a stem cell-specific cytokinesis program is engaged in PHASE ONE through formation of a secondary F-actin ring. That ring is retained for ~8 hours until finally disassembled in PHASE TWO. Only in PHASE TWO does the intercellular bridge between the GSC and its daughter elongate, thin and eventually sever through abscission.
OUR MODEL:
DELAYED GSC ABSCISSION PROMOTES COORDINATED RELEASE OF STEM CELL DAUGHTERS FROM THE NICHE
Our data suggests that delayed GSC abscission serves to coordinate release of stem cell daughters from the niche. When the gonialblast daughter is produced it is not properly encysted but it is ALSO not released from the niche. It is retained in close proximity to the dividing somatic stem cells so that over time, as those cells produce cyst cell daughters, the gonialblast can become properly encysted prior to completion of cytokinesis and release of the entire three cell grouping from the niche. We have found this process to be controlled at two discrete steps:
a PAUSE controlled by signals from the niche and a TRIGGER for abscission controlled by the somatic stem cells.
MECHANISMS CONTROLLING GSC CYTOKINESIS: THE NICHE AND SOMATIC STEM CELLS
JAK/STAT SIGNALING FROM THE NICHE REGULATES THE ABSCISSION PAUSE
The hub cells of the niche produce a number of signals required for self-renewal of the resident stem cells. One main pathway activated in GSCs is Jak/STAT. We have found that STAT is essential to control the abscission PAUSE in GSCs. Loss of STAT leads to repeated abscission failure and formation of "stem cysts"--multiple, interconnected GSC-daughters all associated with the hub.
We find that STAT controls the length of the abscission pause. When STAT is depleted from GSCs, the pause is significantly longer than in controls. As a consequence, there is simply not enough time for abscission to complete before a new round of mitosis initiates. This leads to almost complete failure of abscission in GSCs expressing a STAT RNAi.
SOMATIC STEM CELLS TRIGGER GSC ABSCISSION
Contact between somatic stem cells and GSCs is essential to trigger abscission in GSCs. When encystment is disrupted--somatic stem cells are still present, just no longer touching the GSCs--abscission consistently fails. HOWEVER, the abscission pause is NOT disrupted. Thus, we believe a signal coming from somatic stem cells and acting over a short distance is necessary to trigger the final steps of abscission in GSCs.
CURRENT RESEARCH PROJECTS IN THE LAB
IDENTIFYING THE MECHANISM BY WHICH NICHE JAK/STAT SIGNALING TEMPORALLY CONTROLS THE PAUSE
IDENTIFYING THE SOMATIC TRIGGER OF GSC CYTOKINESIS AND LIVE IMAGING THE COORDINATED RELEASE OF DAUGHTER CELLS FROM THE NICHE
UNDERSTANDING HOW AGING ALTERS STEM CELL INTERACTIONS INCLUDING CYTOKINESIS
MODIFIED CYTOKINESIS AS A CONSERVED NODE OF NICHE CONTROL OVER STEM CELL BEHAVIOR
PUBLISHED WORK
November 2024
CONVERSION OF SOMATIC SEX IDENTITY IN THE TESTIS INDUCES FEMALE-SPECIFIC CELLULAR BEHAVIORS IN THE SOMA AND EARLY OOCYTE SPECIFICATION IN THE GERMLINE
Roach and Lenhart. BioRiv
October 2023
MATING-INDUCED ECDYSONE IN THE TESTIS DISRUPTS SOMA-GERMLINE CONTACTS AND STEM CELL CYTOKINESIS
Roach TV, Lenhart KF. BioRxiv.
January 2019
DIMINISHED JAK/STAT SIGNALING CAUSES EARLY-ONSET AGING DEFECTS IN STEM CELL CYTOKINESIS
Lenhart, KF, Capozzoli, B, Warrick, GSG, DiNardo S. Current Biology: 29 (2). 256-267.
July 2015
SOMATIC CELL ENCYSTMENT PROMOTES ABSCISSION IN GERMLINE STEM CELLS FOLLOWING A REGULATED BLOCK IN CYTOKINESIS
Lenhart KF and DiNardo S. Developmental Cell.