Essential role of mitochondrially encoded large rRNA for germ-line formation in Drosophila embryos

Abstract

In Drosophila, pole cells, the progenitors of the germ line, are induced by the factors localized in the posterior pole region of oocytes and cleavage embryos, or germ plasm. Polar granules in germ plasm are electron-dense structures and have been proposed to contain factors essential for pole cell formation. Mitochondrially encoded large ribosomal RNA (mtlrRNA) has been identified as a component of polar granules. We previously have shown that mtlrRNA is able to rescue embryos that fail to form pole cells as a result of UV irradiation. However, there is a possibility that the function of mtlrRNA is limited to UV-irradiated embryos, and the question of whether mtlrRNA is required for the normal pathway leading to pole cell formation remains unanswered. In this study, we report that the reduction of mtlrRNA in germ plasm by injecting anti-mtlrRNA ribozymes into cleavage embryos leads to their inability to form pole cells. Other components of germ plasm, namely oskar mRNA, germ cell-less mRNA, and Vasa and Tudor proteins appear to be unaffected in these ribozyme-injected embryos. These results support an essential role for mtlrRNA in pole cell formation. We propose that mitochondrially encoded molecules participate in a key event in early cell-type specification.

Figure 1

Effect of ribozymes on the distribution of mtlrRNA and polar plasm components in the late-cleavage embryos at around 60 min after egg laying. In situ hybridization of the mtlrRNA cDNA probe to a control embryo injected with DW (A) and an embryo injected with RbzJ and RbzK (B). Note that mtlrRNA signal in polar plasm completely disappears in a ribozyme-injected embryo (B). Arrow in A shows mtlrRNA signal. As an internal control for in situ hybridization, the embryos also were hybridized with a probe detecting bcd mRNA that localizes at the anterior pole region of early cleavage embryos (39, 40). Arrowheads in A-C indicate bcd mRNA signal. In situ hybridization of osk cDNA probe (C) and gcl cDNA probe (D) to the ribozyme-injected embryos. Arrow in C shows osk mRNA signal. Immunostaining of VAS (E) and TUD (F) in the ribozyme-injected embryos. osk mRNA, gcl mRNA, VAS, and TUD normally accumulated in polar plasm of the ribozyme-injected embryos. Lateral views of the late cleavage embryos are shown; anterior is to left.

Figure 2

Effect of ribozymes on pole cell formation. Posterior pole region of embryos at the cellular blastoderm stage (3 hr after egg laying) are shown. (A) A noninjected embryo. (B) A control embryo injected with DW. (C) An embryo injected with RbzJ and RbzK. In the embryo injected with RbzJ and RbzK, pole cells are completely missing, whereas somatic cell layer appears essentially normal. Arrowheads in A and B indicate pole cells.