maternally inherited piRNAs
In hybrid dysgenesis (Drosophila melanogaster), the progeny of intercrosses between wild males and laboratory-strain females are sterile because of defects in gamete formation. This has something to do with the mobilization of transposons. Depending on the parent of transposon origin, there exist cytoplasmically inherited determinants of the outcome phenotype. These are often transmitted through the maternal germ line.
Networks of small RNAs act through RNA-interference (RNAi) pathways to restrain the spread of selfish genetic elements, regulate gene expression, and many other functions. Like microRNAs and small interfering RNAs, such small species guide Argonaute proteins to silencing targets. The control of mobile elements in germ cells depends on a system composed of Piwi-family proteins (Piwi, Aubergine, and AGO3) and piRNAs. Piwi and Aubergine (Aub) transfer maternal piRNAs into the germ line.
Do maternally deposited small RNAs affect transposon suppression in a heritable fashion? Are piRNAs the maternal suppressor of hybrid dysgenesis?
Julius Brennecke and colleagues recently reported an epigenetic role for maternally inherited piRNAs in transpososon silencing. They found that small RNAs themselves serve as vectors for epigenetic information. “In both P- and I-element–mediated hybrid dysgenesis models, daughters show a markedly different content of Piwi-interacting RNAs (piRNAs) targeting each element, depending on their parents of origin. Such differences persist from fertilization through adulthood. This indicates that maternally deposited piRNAs are important for mounting an effective silencing response and that a lack of maternal piRNA inheritance underlies hybrid dysgenesis.”
Science 28 November 2008:
Vol. 322. no. 5906, pp. 1387 – 1392
DOI: 10.1126/science.116517
Fig. 2. I-R hybrid dysgenesis correlates with maternal piRNA inheritance. (A) Normalized piRNA counts for Repbase transposons are plotted for w1118 inducer and wK reactive ovaries. (B) (Left) Fold differences in piRNA counts comparing w1118 and wK mothers are shown (red line indicates a 1:1 ratio). (Right) Transposon piRNA ratios for mothers, embryos, and F1 progeny (SF: RSF ratio) are shown as a heat map. (C) Scatter plots indicating transposon piRNA correlations between w1118 and wK mothers (top) and their respective intercross progeny (bottom).
synaptic proteins, IGF signaling, and lifespan
In a paper by QueeLim Ch’ng and colleagues, a genetic analysis of presynaptic structure in the worm Caenorhabditis elegans was described after measuring in vivo changes in the distribution of fluorescently tagged presynaptic proteins. Because changes in the abundance of presynaptic proteins often indicate different synaptic functional states, they developed a genetic strategy to systematically analyze protein localization at presynaptic structures by GFP-tagging. 25 mutants were observed showing different aspects of neurotransmission. Global analysis of these data identified novel relationships between particular presynaptic components. Moreover, they also identified several genes that regulate secretion of insulin-like growth factors (IGFs) from neurons, and showed that these genes regulate lifespan, a physiological function of IGF signaling.
Ch’ng Q, Sieburth D, Kaplan JM (2008) Profiling Synaptic Proteins Identifies Regulators of Insulin Secretion and Lifespan. PLoS Genet 4(11): e1000283. doi:10.1371/journal.pgen.1000283
Figure 6. Analysis of DCV accumulation in axons and insulin/IGF secretion. (A) XY plot comparing changes in the punctal fluorescence of SNB-1 synaptobrevin and INS-22 insulin/IGF. Two mutants with more prominent increases in INS-22 insulin/IGF than SNB-1 synaptobrevin are shown as solid orange circles. (B) Secreted INS-22 insulin/IGF expressed in motorneurons accumulates in coelomocytes. (C) Images of INS-22 insulin/IGF accumulation in coelomocytes in wild type and mutant animals. Scale bar, 5 µm. Below is shown the quantification of coelomocyte INS-22 insulin/IGF fluorescence. (D) Images of axonal INS-22 insulin/IGF in wild type animals and mutants with altered INS-22 insulin/IGF coelomocyte fluorescence. Scale bar, 5 µm. Below is shown the quantification of INS-22 insulin/IGF punctal fluorescence. In (C–D), * indicates p<0.05, ** indicates p<0.01, (Student’s T-Test compared to wild type). All error bars are ±SEM. Separate charts indicate data from separate sets of experiments.
Figure 7. Insulin/IGF secretion mutants show lifespan phenotypes. (A–F) Survival curves with indicated genotypes. * indicates significantly different lifespan from wild type (p<0.0001), † indicates significant suppression by daf-16 FOXO (p<0.0001), § indicates significant suppression by daf-2 InsR (p<0.0001), (Log Rank Test).
telomere loss produces genomic instability
Simon Titen and Kent Golic studied telomere loss by breakage of an induced dicentric chromosome in the fruitfly Drosophila melanogaster [Genetics 2008 Dec;180(4):1821-32; Epub 2008 Oct 9]. They found that one outcome of this is cell death through Chk2 and Chk1 controlled p53-dependent apoptotic pathways. Yet they also observed escape from apoptosis and repeated division in some cells that have lost a telomere. This evasion of apoptosis is accompanied by abnormalites such as chromosome fusions, anaphase bridges, aneuploidy, and polyploidy. They also found evidence of bridge–breakage–fusion cycles and chromosome segments without centromeres. They further noted that cells manifesting these signs of genomic instability were much more frequent when the apoptotic mechanisms were crippled. In the end, they concluded that the loss of a single telomere is sufficient to generate genomic instability involving multiple chromosomes and aneuploidy.
They used the FLP recombinase system to examine the cellular responses to loss of a single telomere. This generates a chromosome end lacking a telomere. Synthesis of FLP recombinase is induced by heat shock, which causes recombination between inverted FRTs on sister chromatids to generate dicentric and acentric chromosomes. They believe their strategy closely mimics the situation faced by a mammalian cell in which a single telomere becomes critically short and dysfunctional due to incomplete replication in the absence of telomerase.
Figure 1. Dicentric/acentric chromosome production and segregation. FLP-induced recombination between oppositely oriented FRTs on sister chromatids produces a dicentric chromosome and an acentric chromosome. At anaphase the dicentric chromosome is stretched between the poles and usually breaks. Centromeres are indicated as filled circles, telomeres as filled squares, FRTs as arrows.
Holliday junction resolvases
The formation of four-way DNA junctions, Holliday junctions, account for products formed during meiotic recombination. These junctions are involved recombinational repair of DNA double-stranded breaks. Resolvases are small homodimeric enzymes that resolve Holliday junctions by endonucleolytic cleavage. Resolution is achieved by the introduction of symmetrically related nicks in two strands of like polarity. In a recent study by Stephen Ip and colleagues, nucleases that promote Holliday junction resolution, in a similar manner to Escherichia coli Holliday junction resolvase RuvC, were identified in human cells and also the budding yeast Saccharomyces cerevisiae.
They stated that the identification of GEN1 and Yen1 Holliday junction resolvases suggests that the RuvC model is universal. GEN1 and Yen1 promote Holliday junction resolution by a mechanism analogous to that shown by RuvC, Cce1, Hjc and the bacteriophage resolvases, but they neither do show sequence nor structure conservation with these proteins, instead they belong to the Rad2/XPG family of nucleases. There are three known classes in this group. Class I (XPG in humans and Rad2 in yeast) — endonucleolytic incision at the 3′ side of bubble structures formed during nucleotide excision repair. Class II (FEN1 in humans and Rad27 in yeast) — 5′-flap endonuclease activities for Okazaki fragment processing during DNA replication. Class III — EXO1-related proteins which exhibit 5′–3′ exonuclease activities required for DNA replication, DNA repair and meiotic recombination.
The results described in their study indicate the eukaryotic Holliday junction resolvases comprise Class IV.
Nature. 2008 Nov 20;456(7220):357-61.
Drosophila CG6539 is orthologue of vertebrate gemin3
Mutations in the survival motor neuron (SMN1) gene cause spinal muscular atrophy (SMA), an autosomal recessive disorder characterised by degeneration of spinal cord motor neurons leading to progressive muscular weakness. SMN1 encodes an RNA-binding protein, SMN, which is complexed with Gemin proteins. Vertebrate Gemin3 is the only RNA helicase in the SMN complex. In Drosophila melanogaster, CG6539 was identified as the orthologue of vertebrate gemin3. It was shown to physically interact with SMN, and its loss results into larval/prepupal lethality. Before death, gemin3 mutant larvae exhibit declined mobility and expanded neuromuscular junctions.
Cauchi RJ, Davies KE, Liu J-L (2008) A Motor Function for the DEAD-Box RNA Helicase, Gemin3, in Drosophila. PLoS Genet 4(11): e1000265. doi:10.1371/journal.pgen.1000265
doi:10.1371/journal.pgen.1000265.g008
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perinuclear tethering promotes rDNA repeat stability
Karim Mekhail and colleagues studied a network in the the budding yeast Saccharomyces cerevisiae that stabilizes ribosomal DNA repeats thru interactions between rDNA-associated silencing proteins and proteins of the inner nuclear membrane (INM).
They reported that deletion of either the INM or silencing proteins:
—reduces perinuclear rDNA positioning
—disrupts the nucleolus–nucleoplasm boundary
—induces the formation of recombination foci
—destabilizes the rDNA repeats
“In addition, artificial targeting of rDNA repeats to the INM suppresses the instability observed in cells lacking an rDNA-associated silencing protein that is typically required for peripheral tethering of the repeats.
Moreover, in contrast to Sir2 and its associated nucleolar factors, the INM proteins are not required for rDNA silencing, indicating that Sir2-dependent silencing is not sufficient to inhibit recombination within the rDNA locus.
These findings demonstrate a role for INM proteins in the perinuclear localization of chromosomes and show that tethering to the nuclear periphery is required for the stability of rDNA repeats.”
Their results suggest that Sir2-dependent silencing alone cannot inhibit recombination within the repetitive rDNA locus and that INM-mediated perinuclear chromosome tethering ensures repeat stability.
They expect that the proteins studied are members of perinuclear networks that control recombination at multiple loci to maintain genome stability.
Hutchinson-Guildford progeria syndrome in Drosophila
In humans, Hutchinson-Guildford progeria syndrome is caused by mutations in the LMNA gene. In Drosophila, there are two lamin genes: lamC and Dm0. Previous studies have shown that Drosophila and vertebrate lamin genes share a common ancestor. Drosophila lamin null mutations lead to lethality (either pupal lethality or complete pre-metamorphosis lethalit, depending on which type of lamin). A recent study by Gemma Beard and colleagues looked at ectopic expression of progerin and human lamin A in Drosophila.
… to be continued …
WRN and MUS81
There is a Journal of Cell Biology paper by Annapaola Franchitto and colleagues entitled:
Below is tha abstract:
Failure to stabilize and properly process stalled replication forks results in chromosome instability, which is a hallmark of cancer cells and several human genetic conditions that are characterized by cancer predisposition. Loss of WRN, a RecQ-like enzyme mutated in the cancer-prone disease Werner syndrome (WS), leads to rapid accumulation of double-strand breaks (DSBs) and proliferating cell nuclear antigen removal from chromatin upon DNA replication arrest. Knockdown of the MUS81 endonuclease in WRN-deficient cells completely prevents the accumulation of DSBs after fork stalling. Also, MUS81 knockdown in WS cells results in reduced chromatin recruitment of recombination enzymes, decreased yield of sister chromatid exchanges, and reduced survival after replication arrest. Thus, we provide novel evidence that WRN is required to avoid accumulation of DSBs and fork collapse after replication perturbation, and that prompt MUS81-dependent generation of DSBs is instrumental for recovery from hydroxyurea-mediated replication arrest under such pathological conditions.
Published online October 13, 2008
doi:10.1083/jcb.200803173
The Journal of Cell Biology, Vol. 183, No. 2, 241-252
