Binucleate germ cells in Caenorhabditis elegans are removed by physiological apoptosis

Abstract

Cell death plays a major role during C. elegans oogenesis, where over half of the oogenic germ cells die in a process termed physiological apoptosis. How germ cells are selected for physiological apoptosis, or instead become oocytes, is not understood. Most oocytes produce viable embryos when apoptosis is blocked, suggesting that physiological apoptosis does not function to cull defective germ cells. Instead, cells targeted for apoptosis may function as nurse cells; the germline is syncytial, and all germ cells appear to contribute cytoplasm to developing oocytes. C. elegans has been a leading model for the genetics and molecular biology of apoptosis and phagocytosis, but comparatively few studies have examined the cell biology of apoptotic cells. We used live imaging to identify and examine pre-apoptotic germ cells in the adult gonad. After initiating apoptosis, germ cells selectively export their mitochondria into the shared pool of syncytial cytoplasm; this transport appears to use the microtubule motor kinesin. The apoptotic cells then shrink as they expel most of their remaining cytoplasm, and close off from the syncytium. Shortly thereafter the apoptotic cells restructure their microtubule and actin cytoskeletons, possibly to maintain cell integrity; the microtubules form a novel, cortical array of stabilized microtubules, and actin and cofilin organize into giant cofilin-actin rods. We discovered that some apoptotic germ cells are binucleate; the binucleate germ cells can develop into binucleate oocytes in apoptosis-defective strains, and appear capable of producing triploid offspring. Our results suggest that the nuclear layer of the germline syncytium becomes folded during mitosis and growth, and that binucleate cells arise as the layer unfolds or everts; all of the binucleate cells are subsequently removed by apoptosis. These results show that physiological apoptosis targets at least two distinct populations of germ cells, and that the apoptosis machinery efficiently recognizes cells with two nuclei.

Author summary

Many germ cells die by apoptosis during the development of animal oocytes, including more than half of all germ cells in the model system C. elegans. How individual germ cells are selected for apoptosis, or survival, is not known. Here we study the cell biology of apoptosis. The C. elegans gonad is a syncytium, with nearly 1000 germ “cells” connected to a shared, core cytoplasm. Once apoptosis is initiated, germ cells selectively transport their mitochondria into the gonad core, apparently using the microtubule motor protein kinesin. The apoptotic cells next constrict, expelling most of their remaining cytoplasm into the core, and close off from the gonad core. The microtubule and actin cytoskeletons are remodeled and stabilized, presumably to maintain the integrity of the dying cell. The apoptotic cells form giant cofilin-actin rods, similar to rods described in stressed cultured cells and in human myopathies and neuropathies such as Alzheimer’s and Huntington’s disease. We show that some germ cells are binucleate; these cells appear to form during germline morphogenesis, and are removed by apoptosis. These results demonstrate heterogeneity between oogenic germ cells, and show that the apoptosis machinery efficiently recognizes and removes cells with two nuclei.