Ras and Neuronal Survival
By generating transgenic mice which selectively express a mutant Ras gene in their neurons, Heumann et al. (page 1537) have found that the Ras signaling pathway can trigger neuroprotective mechanisms in adult mice. The findings suggest a novel approach for stabilizing isolated neurons, and may lead to the development of a new class of therapies for neurodegenerative diseases.
Earlier studies suggested that Ras may mediate some neuronal survival signals, but experiments with different types of neurons have given conflicting results. To address the issue in vivo, the authors of the new study created transgenic mice in which the neuronal promoter for the synapsin-1 gene drives expression of constitutively activated Ha-Ras. Neuronal Ras in these mice is constitutively active, leading to phosphorylation of mitogen-activated kinase. Choline acetyltransferase and tyrosine hydroxylase activities, as well as neuropeptide Y expression, are also increased in the transgenic system. While the mice appear healthy and have a normal life span, they exhibit an increase in brain volume caused by cell soma hypertrophy.
The animals were completely protected from motor neuron degeneration after facial nerve lesion, and were significantly protected from neurotoxin-induced degeneration of dopaminergic substantia nigra neurons, a model for Parkinson's disease. Based on these results, the authors suggest that neuronal Ras activation may prove useful in stabilizing donor neurons and treating neurodegenerative diseases.
Role for Wee1 in Apoptosis
Using cell-free extracts from Xenopus eggs to study apoptotic signaling pathways, Smith et al. (page 1391) have discovered a new role for the well-known cell cycle regulator Wee1: the protein appears to play a key role in triggering the apoptotic machinery. Based on their results, Smith et al. propose that Wee1 and the adaptor protein Crk function in a novel apoptotic pathway, and that this activity of Wee1 may be independent of its role in cell cycle regulation.
Previous studies by Smith et al. demonstrated that Crk is required for in vitro apoptosis in the Xenopus egg extract system, and that the isolated SH2 domain of Crk could inhibit apoptosis in the system, presumably by titrating another component of the apoptotic pathway. In the new work, the researchers identified the critical Crk SH2-interacting protein as Wee1. Though Wee1 is known to inhibit Cdc2, its apoptotic signaling is not Cdc2-dependent. The Wee1-mediated apoptotic signal does, however, depend on the presence of Crk in the extracts, and depleting Wee1 from the system significantly delays the onset of apoptosis. The authors speculate that the dual function of Wee1 in frog eggs may make it a key determinant in the decision to proceed with embryogenesis or to undergo apoptosis, and that a similar decision, between checkpoint-mediated cell cycle arrest or Wee1-regulated apoptosis, might take place in other cell types.
Evidence for a Spindle Matrix
Walker et al. (page 1401) cloned and characterized a novel Drosophila nuclear protein, which they propose is part of a complex forming a spindle matrix. Though a spindle matrix had been hypothesized to play a role in organizing the microtubule spindle during mitosis, the new work provides the first direct evidence that a complete spindle matrix forms within the nucleus before microtubule spindle formation.
Walker et al. used a monoclonal antibody to identify a novel protein that is localized in a cell cycle–dependent manner. The 81-kD protein, named skeletor, associates with chromosomes at interphase, but redistributes into a spindle-like structure during prophase, before microtubule spindle formation. The skeletor-containing spindle coaligns with microtubule spindles during metaphase, and persists even when microtubules are depolymerized by nocodazole or low temperature.
Based on their results, the authors propose a model in which chromosome condensation is accompanied by skeletor redistribution, leading to the formation of a spindle structure from a skeletor-containing macromolecular complex at late prophase. In metaphase, microtubule spindle fibers could then coalign with the skeletor-defined spindle with chromosomes positioned at the metaphase plate. The skeletor spindle would then remain intact through anaphase, and as chromosomes decondense in telophase they would reassociate with skeletor. The team is now working to isolate Drosophila mutants with defects in skeletor to further illuminate the function of the spindle matrix.
53BP1 at Sites of Double-strand Breaks
Based on the intracellular localization of 53BP1 after DNA damage, Schultz et al. (page 1381) conclude that the protein participates in the cellular response to DNA double-strand breaks (DSBs). The rapid relocalization of 53BP1, which has previously been hypothesized to act as a transcriptional coactivator of the p53 tumor suppressor, suggests that 53BP1 is an early participant in the DSB response.
Because it contains domains with high homology to the Saccharomyces cerevisiae Rad9p DNA damage checkpoint protein, Schultz et al. examined the localization of 53BP1 in cells before and after exposure to ionizing radiation. Before irradiation, the protein exhibits diffuse nuclear staining, but it localizes to discreet nuclear foci within 5–15 min after irradiation. The foci were only induced by agents that cause DSBs, and their number corresponds to the number of DSBs, decreasing over time as DNA repair is carried out. Wortmannin, which slows DSB repair, also slowed the formation of 53BP1 foci. Interestingly, 53BP1-containing foci colocalize with H2AX and Mre11/NBS1/Rad50 foci, which are associated with sites of DSB processing. The researchers propose that phosphorylation of H2AX may lead to the recruitment of 53BP1 and other proteins involved in DNA damage repair.
Histone H2B Variant in Sperm Telomeres
Beginning on page 1591, Gineitis et al. describe the isolation and partial purification of a telomere-binding complex from human sperm. Their analysis shows that the complex, called hSTBP, does not contain the known somatic telomere proteins TRF1, TRF2, and Ku, but does contain a sperm-specific variant of histone H2B. The localization of the complex suggests that hSTBP plays a role in the membrane attachment of telomeres in sperm.
Earlier studies had demonstrated that the telomeres in germ-line cells are structurally and biochemically different from those in somatic cells, but the protein complexes associated with sperm telomeres remained poorly characterized. In the new work, Gineitis et al. isolated a detergent-soluble protein complex that interacts with double-stranded telomeric DNA in sperm. Though the complex lacks known somatic-cell telomere-binding proteins, it contains spH2B, a sperm-specific variant histone that is biochemically distinct from the major replication-dependent H2B. Foci of spH2B in human sperm nuclei partially colocalize with telomere DNA, and in vitro binding experiments suggest that spH2B is involved in the DNA recognition activity of hSTBP. The authors hypothesize that hSTBP-mediated membrane attachment of telomeres may play a role in chromosome withdrawal after fertilization.
Apicoplasts Hold On
The Apicomplexa are intracellular protozoan parasites that cause diseases such as malaria and toxoplasmosis. Despite their clear eukaryotic nature, many Apicomplexa are unexpectedly sensitive to prokaryotic-specific antibiotics. The explanation appears to be the apicoplast, a strange organelle containing a 35-kb episome most closely related to chloroplast organellar genomes. The apicoplast is thought to have started life as a free-living cyanobacterium, before primary endosymbiosis yielded a chloroplast resident in an algal cell, and secondary endosymbiosis of the algal cell resulted in a vestigial (but nonetheless essential) organelle surrounded by four membranes.
The bizarre behavior did not stop there. The parent Apicomplexa cells, such as the Toxoplasma gondii cells studied by Striepen et al. on page 1423, segregate their replicated DNA within an intact nucleus, with the daughter nuclei budding from this nucleus driven by the intranuclear spindle. Mother cells can produce either two or many more daughter nuclei before daughter cells are formed, so making sure that other cellular components segregate with all these daughter nuclei is quite a task. Striepen et al. report that T. gondii's solution is to link the apicoplast to the centrosome of each daughter nucleus. They observe that apicoplast and nuclear division are synchronized (unlike the division of plant nuclear and plastid genomes), and the apicoplast and centrosome are closely apposed throughout the cell cycle.
In addition to keeping one apicoplast with each nucleus, the centrosome connection may transduce the force of the expanding mitotic spindle to pull one apicoplast into two. Consistent with this theory, no homologs of the FtsZ-like proteins that have been implicated in the fission of other plastids have been identified in Apicomplexa thus far.
Alan W. Dove, 350 E. Willow Grove Ave. #406, Philadelphia, PA firstname.lastname@example.org