Nucleolar proteins and ultrastructure in bovine in vivo developed, in vitro produced, and parthenogenetic cleavage-stage embryos

In the present study, ribosomal RNA (rRNA) gene activation, monitored through nucleolus development, was studied by autoradiography following 3H‐uridine incubation, transmission electron microscopy, and immunofluorescence confocal laser scanning microscopy of key nucleolar proteins involved in rRNA transcription (topoisomerase I, upstream binding factor, and RNA polymerase I) and processing (fibrillarin, nucleolin, and nucleophosmin) in in vivo developed, in vitro produced, and parthenogenetic bovine embryos. In general, in vivo developed embryos displayed formation of fibrillo‐granular nucleoli during the 4th post‐fertilization cell cycle. During the previous stages of development, nucleolus precursor bodies (NPBs) were observed. However, on some occasions the initial steps of nucleolus formation were observed already at the 2‐ and 4‐cell stage in cases where such embryos were collected from superovulated animals together with later embryonic stages presenting nucleolar development and autoradiographic labeling. The in vitro produced embryos displayed very synchronous formation of fibrillo‐granular nucleoli and autoradiographic labeling during the 4th cell cycle. In vivo developed and in vitro produced embryos displayed allocation of nucleolar proteins to fibrillar and granular compartments of the developing nucleoli during the 4th cell cycle. The parthenogenetic embryos typically displayed formation of fibrillo‐ granular nucleoli during the 5th cell cycle and autoradiographic labeling was not observed until the morula stage. Moreover, the 1‐, 2‐, and 4‐cell parthenogenetic embryos practically lacked NPBs. On the other hand, parthenogenetic embryos displayed allocation of nucleoar proteins to nuclear entities during the 4th cell cycle. In conclusion, both in vivo developed and in vitro produced bovine embryos displayed activation of transcription and nucleolar development during the 4th cell cycle. However, in vivo developed embryos flushed together with later developmental stages displayed premature activation of these processes. Parthenogenetic bovine embryos, on the other hand, displayed a delayed activation. Mol. Reprod. Dev. 65: 73–85, 2003. © 2003 Wiley‐Liss, Inc.