Differences in protein mobility between pioneer versus follower growth cones

Navigating growth cones need to integrate, process and respond to guidance signals, requiring dynamic information transfer within and between different compartments. Studies have shown that, faced with different navigation challenges, growth cones display dynamic changes in growth kinetics and morphologies. However, it remains unknown whether these are paralleled by differences in their internal molecular dynamics. To examine whether there are protein mobility differences during guidance, we developed multiphoton fluorescence recovery after photobleaching methods to determine molecular diffusion rates in pathfinding growth cones in vivo. Actively navigating growth cones (leaders) have consistently longer recovery times than growth cones that are fasciculated and less actively navigating (followers). Pharmacological perturbations of the cytoskeleton point to actin as the primary modulator of diffusion in differently behaving growth cones. This approach provides a powerful means to quantify mobility of specific proteins in neurons in vivo and reveals that diffusion is important during axon navigation

Intrabody-mediated diverting of HP1β to the cytoplasm induces co-aggregation of H3-H4 histones and lamin-B receptor

Diverting a protein from its intracellular location is a unique property of intrabodies. To interfere with the intracellular traffic of heterochromatin protein 1β (HP1β) in living cells, we have generated a cytoplasmic targeted anti-HP1β intrabody, specifically directed against the C-terminal portion of the molecule. HP1β is a conserved component of mouse and human constitutive heterochromatin involved in diverse nuclear functions including gene silencing, DNA repair and nuclear membrane assembly. We found that the anti-HP1β intrabody sequesters HP1β into cytoplasmic aggregates, inhibiting its traffic to the nucleus. Lamin B receptor (LBR) and a subset of core histones (H3/H4) are also specifically co-sequestered in the cytoplasm of anti-HP1β intrabody-expressing cells. Methylated histone H3 at K9 (Me9H3), a marker of constitutive heterochromatin, is not affected by the anti-HP1β intrabody expression. Hyper-acetylating conditions completely dislodge H3 from HP1β:LBR containing aggregates. The expression of anti-HP1β scFv fragments induces apoptosis, associated with an alteration of nuclear morphology. Both these phenotypes are specifically rescued either by overexpression of recombinant full length HP1β or by HP1β mutant containing the chromoshadow domain, but not by recombinant LBR protein. The HP1β-chromodomain mutant, on the other hand, does not rescue the phenotypes, but does compete with LBR for binding to HP1β. These findings provide new insights into the mode of action of cytoplasmic-targeted intrabodies and the interaction between HP1β and its binding partners involved in peripheral heterochromatin organisation

Heterochromatin Dynamics

Heterochromatin is usually thought of as a stable and inactive region of the genome. Not so, according to a study published earlier this yea

Microbiome turnover during offspring development varies with maternal care, but not moult, in a hemimetabolous insect

The ecological success of insects often depends on their association with beneficial microbes. However, insect development involves repeated moults, which can have dramatic effects on their microbial communities. Here, we investigated whether and how moulting affects the microbiome of a hemimetabolous insect, and whether maternal care can modulate these effects. We reared European earwig juveniles with or without mothers and used 16S rRNA metabarcoding to analyse the prokaryotic fraction of the core microbiome of eggs, recently and old moulted individuals at four developmental stages and the resulting adults. The 218 samples obtained showed that the microbiome diversity changed non-linearly during development and that these changes were associated with bacterial biomarkers. Surprisingly, these changes did not occur during moulting, but rather between the beginning and end of certain developmental stages. We also found that access to maternal care affected the microbiome of both juveniles and adults, even when the last contact with mothers was two months before adulthood. Overall, these results provide new insights into our understanding of the (in)stability of the prokaryotic microbiome in hemimetabolous insects and its independence from moult. More generally, they question the role of microbiome acquisition through maternal care in maintaining family life in species where this behaviour is facultative

Long non-coding RNAs: spatial amplifiers that control nuclear structure and gene expression

Over the past decade, it has become clear that mammalian genomes encode thousands of long non-coding RNAs (lncRNAs), many of which are now implicated in diverse biological processes. Recent work studying the molecular mechanisms of several key examples — including Xist, which orchestrates X chromosome inactivation — has provided new insights into how lncRNAs can control cellular functions by acting in the nucleus. Here we discuss emerging mechanistic insights into how lncRNAs can regulate gene expression by coordinating regulatory proteins, localizing to target loci and shaping three-dimensional (3D) nuclear organization. We explore these principles to highlight biological challenges in gene regulation, in which lncRNAs are well-suited to perform roles that cannot be carried out by DNA elements or protein regulators alone, such as acting as spatial amplifiers of regulatory signals in the nucleus

From the cradle to the grave:Green turtle hatchlings (Chelonia mydas) preyed upon by two-spots red snappers (Lutjanus bohar)

The observation of trophic interactions such as predation provide valuable information to model food webs and better understand ecosystem functioning. Such information is crucial for rare and endangered species in order to adapt management measures and ensure their conservation. However, trophic interactions are rarely observed in the marine realm, even for well-known or widespread species. During a scientific cruise in the Scattered Islands (Southwestern Indian Ocean), we observed endangered green turtle hatchlings (Chelonia mydas) in the gut content of two subadults two-spots red snappers (Lutjanus bohar). This trophic link involving emblematic species has not been previously described. The two-spots red snapper is a widespread coral reef fish in the tropical Indo-Pacific. Although it is unclear how fish predation affects marine turtle population dynamics, the occurrence of hatchlings in all the snapper samples suggests that fish could be significant sources of predation. Yet this predation pressure remains to be further studied and quantified to be considered in marine turtle population monitoring

Identification and characterization of a novel ubiquitous nucleolar protein ‘NARR’ encoded by a gene overlapping the rab34 oncogene

There are only few reports on protein products originating from overlapping mammalian genes even though computational predictions suggest that an appreciable fraction of mammalian genes could potentially overlap. Mass spectrometry-based proteomics has now acquired the tools to probe proteins in an unbiased manner, providing direct evidence of the output of the genomic and gene expression machinery. In particular, proteomics can refine gene predictions and discover novel gene-processing events and gene arrangements. Here, we report the mass spectrometric discovery and biochemical validation of the novel protein encoded by a gene overlapping rab34 oncogene. The novel protein is highly conserved in mammals. In humans, it contains 13 distinct Nine-Amino acid Residue-Repeats (NARR) with the consensus sequence PRVIV(S/T)PR in which the serine or threonine residues are phosphorylated during M-phase. NARR is ubiquitously expressed and resides in nucleoli where it colocalizes with ribosomal DNA (rDNA) gene clusters. Its distribution only partially overlaps with upstream binding factor, one of the main regulators of RNA Polymerase I activity, and is entirely uncoupled from it in mitotic cells and upon inhibition of transcription. NARR only partially colocalizes with fibrillarin, the pre-ribosomal RNA-processing protein, positioning NARR in a separate niche within the rDNA cluster

Interplay of ribosomal DNA Loci in nucleolar dominance: dominant NORs are up-regulated by chromatin dynamics in the wheat-rye system

Background: Chromatin organizational and topological plasticity, and its functions in gene expression regulation, have been strongly revealed by the analysis of nucleolar dominance in hybrids and polyploids where one parental set of ribosomal RNA (rDNA) genes that are clustered in nucleolar organizing regions (NORs), is rendered silent by epigenetic pathways and heterochromatization. However, information on the behaviour of dominant NORs is very sparse and needed for an integrative knowledge of differential gene transcription levels and chromatin specific domain interactions. Methodology/Principal Findings: Using molecular and cytological approaches in a wheat-rye addition line (wheat genome plus the rye nucleolar chromosome pair 1R), we investigated transcriptional activity and chromatin topology of the wheat dominant NORs in a nucleolar dominance situation. Herein we report dominant NORs up-regulation in the addition line through quantitative real-time PCR and silver-staining technique. Accompanying this modification in wheat rDNA trascription level, we also disclose that perinucleolar knobs of ribosomal chromatin are almost transcriptionally silent due to the residual detection of BrUTP incorporation in these domains, contrary to the marked labelling of intranucleolar condensed rDNA. Further, by comparative confocal analysis of nuclei probed to wheat and rye NORs, we found that in the wheat-rye addition line there is a significant decrease in the number of wheat-origin perinucleolar rDNA knobs, corresponding to a diminution of the rDNA heterochromatic fraction of the dominant (wheat) NORs. Conclusions/Significance: We demonstrate that inter-specific interactions leading to wheat-origin NOR dominance results not only on the silencing of rye origin NOR loci, but dominant NORs are alsomodified in their transcriptional activity and interphase organization. The results show a cross-talk between wheat and rye NORs, mediated by ribosomal chromatin dynamics, revealing a conceptual shift from differential amphiplasty to ‘mutual amphiplasty’ in the nucleolar dominance process.This work was supported by the Fundação para a Ciência e Tecnologia (projects POCI/BIA-BDE/57575/2004 to M.S. and POCI/BIA-BCM/59389/2004 to N.N.

Heterochromatin protein 1 is recruited to various types of DNA damage

Heterochromatin protein 1 (HP1) family members are chromatin-associated proteins involved in transcription, replication, and chromatin organization. We show that HP1 isoforms HP1-α, HP1-β, and HP1-γ are recruited to ultraviolet (UV)-induced DNA damage and double-strand breaks (DSBs) in human cells. This response to DNA damage requires the chromo shadow domain of HP1 and is independent of H3K9 trimethylation and proteins that detect UV damage and DSBs. Loss of HP1 results in high sensitivity to UV light and ionizing radiation in the nematode Caenorhabditis elegans, indicating that HP1 proteins are essential components of DNA damage response (DDR) systems. Analysis of single and double HP1 mutants in nematodes suggests that HP1 homologues have both unique and overlapping functions in the DDR. Our results show that HP1 proteins are important for DNA repair and may function to reorganize chromatin in response to damage