Temporal and Spatial Changes in Phyllosphere Microbiome of Acacia Trees Growing in Arid Environments.

Background: The evolutionary relationships between plants and their microbiomes are of high importance to the survival of plants in general and even more in extreme conditions. Changes in the plant's microbiome can affect plant development, growth, fitness, and health. Along the arid Arava, southern Israel, acacia trees (Acacia raddiana and Acacia tortilis) are considered keystone species. In this study, we investigated the ecological effects of plant species, microclimate, phenology, and seasonality on the epiphytic and endophytic microbiome of acacia trees. One hundred thirty-nine leaf samples were collected throughout the sampling year and were assessed using 16S rDNA gene amplified with five different primers (targeting different gene regions) and sequenced (150 bp paired-end) on an Illumina MiSeq sequencing platform. Results: Epiphytic bacterial diversity indices (Shannon-Wiener, Chao1, Simpson, and observed number of operational taxonomic units) were found to be nearly double compared to endophyte counterparts. Epiphyte and endophyte communities were significantly different from each other in terms of the composition of the microbial associations. Interestingly, the epiphytic bacterial diversity was similar in the two acacia species, but the canopy sides and sample months exhibited different diversity, whereas the endophytic bacterial communities were different in the two acacia species but similar throughout the year. Abiotic factors, such as air temperature and precipitation, were shown to significantly affect both epiphyte and endophytes communities. Bacterial community compositions showed that Firmicutes dominate A. raddiana, and Proteobacteria dominate A. tortilis; these bacterial communities consisted of only a small number of bacterial families, mainly Bacillaceae and Comamonadaceae in the endophyte for A. raddiana and A. tortilis, respectively, and Geodematophilaceae and Micrococcaceae for epiphyte bacterial communities, respectively. Interestingly, ~60% of the obtained bacterial classifications were unclassified below family level (i.e., "new"). Conclusions: These results shed light on the unique desert phyllosphere microbiome highlighting the importance of multiple genotypic and abiotic factors in shaping the epiphytic and endophytic microbial communities. This study also shows that only a few bacterial families dominate both epiphyte and endophyte communities, highlighting the importance of climate change (precipitation, air temperature, and humidity) in affecting arid land ecosystems where acacia trees are considered keystone species.

Neat1 in hematopoietic stem cells.

Hematopoietic Stem Cells (HSCs) generate blood and immune cells through a hierarchical process of differentiation. Genes that regulate this process are of great interest for understanding normal and also malignant hematopoiesis. Surprisingly, however, very little is known about long-non-coding RNAs (lncRNA) in HSCs. Neat1 is a lncRNA that plays a major role in the formation of sub-nuclear structures called paraspeckles, and was reported to regulate proliferation and differentiation in other cells types. We detected Neat1 expression using RNA-seq data and RT-qPCR in HSCs, progenitors and effector immune cells, by specific detection of its isoforms. Neat1 is highly expressed in stem and progenitor cells, yet it shows significant reduction in granulocytes. Microscopically, Neat1 is detected as sharp nuclear foci. Paraspeckle proteins NONO and PSPC1 are detected as aggregated nuclear foci in fresh primary hematopoietic cells, and in cultured cells. Induction of differentiation in vitro was found to enhance Neat1 expression. Taken together, our data demonstrate for the first time the expression of Neat1 and paraspeckles formation in HSCs and along hematopoiesis.

A Differentiation Transcription Factor Establishes Muscle-Specific Proteostasis in Caenorhabditis elegans.

Safeguarding the proteome is central to the health of the cell. In multi-cellular organisms, the composition of the proteome, and by extension, protein-folding requirements, varies between cells. In agreement, chaperone network composition differs between tissues. Here, we ask how chaperone expression is regulated in a cell type-specific manner and whether cellular differentiation affects chaperone expression. Our bioinformatics analyses show that the myogenic transcription factor HLH-1 (MyoD) can bind to the promoters of chaperone genes expressed or required for the folding of muscle proteins. To test this experimentally, we employed HLH-1 myogenic potential to genetically modulate cellular differentiation of Caenorhabditis elegans embryonic cells by ectopically expressing HLH-1 in all cells of the embryo and monitoring chaperone expression. We found that HLH-1-dependent myogenic conversion specifically induced the expression of putative HLH-1-regulated chaperones in differentiating muscle cells. Moreover, disrupting the putative HLH-1-binding sites on ubiquitously expressed daf-21(Hsp90) and muscle-enriched hsp-12.2(sHsp) promoters abolished their myogenic-dependent expression. Disrupting HLH-1 function in muscle cells reduced the expression of putative HLH-1-regulated chaperones and compromised muscle proteostasis during and after embryogenesis. In turn, we found that modulating the expression of muscle chaperones disrupted the folding and assembly of muscle proteins and thus, myogenesis. Moreover, muscle-specific over-expression of the DNAJB6 homolog DNJ-24, a limb-girdle muscular dystrophy-associated chaperone, disrupted the muscle chaperone network and exposed synthetic motility defects. We propose that cellular differentiation could establish a proteostasis network dedicated to the folding and maintenance of the muscle proteome. Such cell-specific proteostasis networks can explain the selective vulnerability that many diseases of protein misfolding exhibit even when the misfolded protein is ubiquitously expressed.

Chaperone families and interactions in metazoa.

Quality control is an essential aspect of cellular function, with protein folding quality control being carried out by molecular chaperones, a diverse group of highly conserved proteins that specifically identify misfolded conformations. Molecular chaperones are thus required to support proteins affected by expressed polymorphisms, mutations, intrinsic errors in gene expression, chronic insult or the acute effects of the environment, all of which contribute to a flux of metastable proteins. In this article, we review the four main chaperone families in metazoans, namely Hsp60 (where Hsp is heat-shock protein), Hsp70, Hsp90 and sHsps (small heat-shock proteins), as well as their co-chaperones. Specifically, we consider the structural and functional characteristics of each family and discuss current models that attempt to explain how chaperones recognize and act together to protect or recover aberrant proteins.

Challenging muscle homeostasis uncovers novel chaperone interactions in Caenorhabditis elegans.

Proteome stability is central to cellular function and the lifespan of an organism. This is apparent in muscle cells, where incorrect folding and assembly of the sarcomere contributes to disease and aging. Apart from the myosin-assembly factor UNC-45, the complete network of chaperones involved in assembly and maintenance of muscle tissue is currently unknown. To identify additional factors required for sarcomere quality control, we performed genetic screens based on suppressed or synthetic motility defects in Caenorhabditis elegans. In addition to ethyl methyl sulfonate-based mutagenesis, we employed RNAi-mediated knockdown of candidate chaperones in unc-45 temperature-sensitive mutants and screened for impaired movement at permissive conditions. This approach confirmed the cooperation between UNC-45 and Hsp90. Moreover, the screens identified three novel co-chaperones, CeHop (STI-1), CeAha1 (C01G10.8) and Cep23 (ZC395.10), required for muscle integrity. The specific identification of Hsp90 and Hsp90 co-chaperones highlights the physiological role of Hsp90 in myosin folding. Our work thus provides a clear example of how a combination of mild perturbations to the proteostasis network can uncover specific quality control modules.

Regulation of cellular protein quality control networks in a multicellular organism.

The long-term health of all metazoan cells is linked to protein quality control, which is achieved by proteostasis, a complex network of molecular interactions that determines the health of the proteome under physiological or stress conditions. Studying the regulation of cellular proteostasis in the context of the whole organism has unraveled multiple layers of cell-nonautonomous regulation, including neuronal regulation, cell-to-cell stress signals and endocrine signaling that affect growth, development and aging. Here, we discuss emerging concepts in cell-nonautonomous regulation of protein quality control networks. The identification of organismal modulators of cellular proteostasis may present novel, yet general targets for misfolding disease intervention.