Mitochondria in eosinophils: Functional role in apoptosis but not respiration

In most eukaryotic cells, mitochondria use the respiratory chain to produce a proton gradient, which is then harnessed for the synthesis of ATP. Recently, mitochondrial roles in regulation of apoptosis have been discovered in many cell types. Eosinophils (Eos) die by apoptosis, but the presence and function of mitochondria in Eos are unknown. This study found that Eos contain mitochondria in small numbers, as shown by labeling with membrane potential-sensitive dyes and in situ PCR for a mitochondrial gene. Eos generate mitochondrial membrane potential from hydrolysis of ATP rather than from respiration, as shown by mitochondrial respiratory inhibitors and mitochondrial uncouplers. The mitochondria provide insignificant respiration but can induce apoptosis, as shown by using the mitochondrial F1F0-ATPase inhibitor oligomycin and translocation of cytochrome c. Thus during differentiation of Eos, although respiration is lost, the other central role of mitochondria, the induction of apoptosis, is retained.

Roles for genomic imprinting and the zygotic genome in placental development

The placenta contains several types of feto-maternal interfaces where zygote-derived cells interact with maternal cells or maternal blood for the promotion of fetal growth and viability. The genetic factors regulating the interactions between different cell types within feto-maternal interfaces and the relative contributions of the maternal and zygotic genomes are poorly understood. Genomic imprinting, the epigenetic process responsible for parental origin-dependent functional differences between homologous chromosomes, has been proposed to contribute to these events. Previous studies showed that mouse conceptuses with an absence of imprinted differences between the two copies of chromosome 12 (upon paternal inheritance of both copies) die late in gestation and have a variety of defects, including placentomegaly. Here we examined the role of chromosome 12 imprinting in these placentae in more detail. We show that the spatial interactions between different cell types within feto-maternal interfaces are defective and identify abnormal behaviors in both zygote-derived and maternal cells that are attributed to the genome of the zygote but not the mother. These include compromised invasion of the maternal decidualized endometrium and the central maternal artery situated within it by zygote-derived trophoblast, abnormalities in the wall of the central maternal artery, and defects within the zygote-derived cellular layer of the labyrinth, which is in direct contact with maternal blood. These findings demonstrate multiple roles for chromosome 12 imprinting in the placenta that have not previously been associated with imprinting effects. They provide insights into the function of imprinting in placental development and have evolutionary and clinical implications.

Characterization of mouse angiogenin-related protein: implications for functional studies on angiogenin.

Angiogenin-related protein (Angrp), the putative product of a recently discovered mouse gene, shares 78% sequence identity with mouse angiogenin (Ang). In the present study, the relationship of Angrp to Ang has been investigated by producing both proteins in bacteria and comparing their functional properties. We find that mouse Ang is potently angiogenic, but Angrp is not, even when assayed at relatively high doses. A deficiency in catalytic capacity, which is essential for the biological activity of Ang, does not appear to underlie Angrp's lack of angiogenicity. In fact, Angrp has somewhat greater ribonucleolytic activity toward tRNA and dinucleotide substrates than does Ang. Instead, an inability to bind cellular receptors is implicated since Angrp does not inhibit Ang-induced angiogenesis. Poor conservation of the Ang receptor recognition sequence 58-69 in Angrp most likely contributes to this defect. However, other substitutions must also influence receptor binding since an Angrp quadruple mutant that is identical to Ang in this segment still lacks both angiogenic activity and the capacity to inhibit Ang. The functional differences between Ang and Angrp, together with evidence presented herein that Angrp is regulated differently than Ang, suggest that the roles of the two proteins in vivo may be quite distinct.

Characterization and functional ordering of Slu7p and Prp17p during the second step of pre-mRNA splicing in yeast.

Temperature-sensitive alleles in four genes (slu7-1, prp16-2, prp17-1, and prp18-1) are known to confer a specific block to the second chemical step of pre-mRNA splicing in vivo in the yeast Saccharomyces cerevisiae. Previous studies showed that Prp16p and Prp18p are required solely for the second step in vitro. The RNA-dependent ATPase, Prp16p, functions at a stage in splicing when ATP is required, whereas Prp18p functions at an ATP-independent stage. Here we use immunodepletion to show that the roles of Slu7p and Prp17p are also confined to the second step of splicing. We find that extracts depleted of Prp17p require both Prp17p and ATP for slicing complementation, whereas extracts depleted of Slu7p require only the addition of Slu7p. These different ATP requirements suggest that Prp16p and Prp17p function before Prp18p and Slu7p. Although SLU7 encodes an essential gene product, we find that a null allele of prp17 is temperature-sensitive for growth and has a partial splicing defect in vitro. Finally, high-copy suppression experiments indicate functional interactions between PRP16 and PRP17, PRP16 and SLU7, and SLU7 and PRP18. Taken together, the results suggest that these four factors may function within a multi-component complex that has both an ATP-dependent and an ATP-independent role in the second step of pre-mRNA splicing.