Comparative analysis of tandem repeats from hundreds of species reveals unique insights into centromere evolution

Background

Centromeres are essential for chromosome segregation, yet their DNA sequences evolve rapidly. In most animals and plants that have been studied, centromeres contain megabase-scale arrays of tandem repeats. Despite their importance, very little is known about the degree to which centromere tandem repeats share common properties between different species across different phyla. We used bioinformatic methods to identify high-copy tandem repeats from 282 species using publicly available genomic sequence and our own data.

Results

Our methods are compatible with all current sequencing technologies. Long Pacific Biosciences sequence reads allowed us to find tandem repeat monomers up to 1,419 bp. We assumed that the most abundant tandem repeat is the centromere DNA, which was true for most species whose centromeres have been previously characterized, suggesting this is a general property of genomes. High-copy centromere tandem repeats were found in almost all animal and plant genomes, but repeat monomers were highly variable in sequence composition and length. Furthermore, phylogenetic analysis of sequence homology showed little evidence of sequence conservation beyond approximately 50 million years of divergence. We find that despite an overall lack of sequence conservation, centromere tandem repeats from diverse species showed similar modes of evolution.

Conclusions

While centromere position in most eukaryotes is epigenetically determined, our results indicate that tandem repeats are highly prevalent at centromeres of both animal and plant genomes. This suggests a functional role for such repeats, perhaps in promoting concerted evolution of centromere DNA across chromosomes.     

Anticancer immunotherapy by CTLA-4 blockade: obligatory contribution of IL-2 receptors and negative prognostic impact of soluble CD25

The cytotoxic T lymphocyte antigen-4 (CTLA-4)-blocking antibody ipilimumab induces immune-mediated long-term control of metastatic melanoma in a fraction of patients. Although ipilimumab undoubtedly exerts its therapeutic effects via immunostimulation, thus far clinically useful, immunologically relevant biomarkers that predict treatment efficiency have been elusive. Here, we show that neutralization of IL-2 or blocking the α and β subunits of the IL-2 receptor (CD25 and CD122, respectively) abolished the antitumor effects and the accompanying improvement of the ratio of intratumoral T effector versus regulatory cells (Tregs), which were otherwise induced by CTLA-4 blockade in preclinical mouse models. CTLA-4 blockade led to the reduction of a suppressive CD4⁺ T cell subset expressing Lag3, ICOS, IL-10 and Egr2 with a concomitant rise in IL-2-producing effector cells that lost FoxP3 expression and accumulated in regressing tumors. While recombinant IL-2 improved the therapeutic efficacy of CTLA-4 blockade, the decoy IL-2 receptor α (IL-2Rα, sCD25) inhibited the anticancer effects of CTLA-4 blockade. In 262 metastatic melanoma patients receiving ipilimumab, baseline serum concentrations of sCD25 represented an independent indicator of overall survival, with high levels predicting resistance to therapy. Altogether, these results unravel a role for IL-2 and IL-2 receptors in the anticancer activity of CTLA-4 blockade. Importantly, our study provides the first immunologically relevant biomarker, namely elevated serum sCD25, that predicts resistance to CTLA-4 blockade in patients with melanoma.     

Supplemental risk evaluations and status of Puccinia carduorum for biological control of musk thistle

Tests of seven rare and endangered native North American Cirsium species and four modern artichoke lines were requested in response to a proposal for introduction of Puccinia carduorum into the United States for biological control of musk thistle (Carduus nutans ssp. leiophyllus). These tests were supplemental to an earlier extensive host-range study that established P. carduorum from musk thistle as host specific, useful for biological control, and suitable for limited field tests in Virginia. Test plants in the current study were evaluated in support of a proposal to use the rust in the western United States, and particularly, in California. None of the test plants in this study had been evaluated in previous assessments and all were either rare, endangered or threatened in California. Tests were conducted in both field and greenhouse settings. Field tests were run for two seasons, and test plants were inoculated by natural spread of the pathogen from source plants inside rings of test plants. Greenhouse tests involved direct inoculation under optimal conditions of dew and temperature (18–20 °C, 16 h) for infection. None of the seven Cirsium species or subspecies tested became infected by P. carduorum, either in field or greenhouse tests, compared to infection of 98% of the individual musk thistle plants (n = 102) from all the studies. Modern artichoke cultivars were tested only by direct inoculation under optimal greenhouse conditions. All artichoke plants (n = 115) either were immune (no macroscopic symptoms, n = 69) or at most, resistant (n = 46); pustules on all but two of the resistant plants were very small (0.30 mm diam). Despite infections on artichokes, P. carduorum could not be maintained on artichokes under optimal greenhouse conditions. These results confirm earlier findings from host-range tests and risk assessments of P. carduorum. This information suggests that rare, threatened, or endangered Cirsium spp. and modern artichoke cultivars are not likely to be adversely affected by the use of P. carduorum for biological control of musk thistle. These data have been reviewed by grower groups and regulatory agencies in a proposal for permission to use the rust for musk thistle control throughout the United States.     

Activation of hypothalamic RIP‐Cre neurons promotes beiging of WAT via sympathetic nervous system

Activation of brown adipose tissue (BAT) and beige fat by cold increases energy expenditure. Although their activation is known to be differentially regulated in part by hypothalamus, the underlying neural pathways and populations remain poorly characterized. Here, we show that activation of rat‐insulin‐promoter‐Cre (RIP‐Cre) neurons in ventromedial hypothalamus (VMH) preferentially promotes recruitment of beige fat via a selective control of sympathetic nervous system (SNS) outflow to subcutaneous white adipose tissue (sWAT), but has no effect on BAT. Genetic ablation of APPL2 in RIP‐Cre neurons diminishes beiging in sWAT without affecting BAT, leading to cold intolerance and obesity in mice. Such defects are reversed by activation of RIP‐Cre neurons, inactivation of VMH AMPK, or treatment with a β3‐adrenergic receptor agonist. Hypothalamic APPL2 enhances neuronal activation in VMH RIP‐Cre neurons and raphe pallidus, thereby eliciting SNS outflow to sWAT and subsequent beiging. These data suggest that beige fat can be selectively activated by VMH RIP‐Cre neurons, in which the APPL2–AMPK signaling axis is crucial for this defending mechanism to cold and obesity.     

Best linear unbiased prediction of host-range of the facultative parasite Colletotrichum gloeosporioides f. sp. salsolae, a potential biological control agent of Russian thistle

Russian thistle or tumbleweed (Salsola tragus L.) is an introduced invasive weed in N. America. It is widely distributed in the US and is a target of biological control efforts.The fungus Colletotrichum gloeosporioides (Penz.) Penz. & Sacc. in Penz. f. sp. salsolae (CGS) is a facultative parasite under evaluation for classical biological control of this weed. Host-range tests were conducted with CGS in quarantine to determine whether the fungus is safe to release in N. America. Ninetytwo accessions were analyzed from 19 families: Aizoaceae, Alliaceae, Amaranthaceae, Apiaceae, Asteraceae, Brassicaceae, Cactaceae, Campanulaceae, Chenopodiaceae, Cucurbitaceae, Cupressaceae, Fabaceae, Malvaceae, Nyctaginaceae, Phytolaccaceae, Poaceae, Polygonaceae, Sarcobataceae, and Solanaceae and 10 tribes within the Chenopodiaceae: Atripliceae, Beteae, Camphorosmeae, Chenopodieae, Corispermeae, Halopepideae, Polycnemeae, Salicornieae, Salsoleae, and Suaedeae. These included 62 genera and 120 species. To facilitate interpretation of results, disease reaction data were combined with a relationship matrix derived from internal transcribed spacer DNA sequences and analyzed with mixed model equations to produce Best Linear Unbiased Predictors (BLUPs) for each species. Twenty-nine species (30 accessions) from seven closely-related Chenopodiaceae tribes had significant levels of disease severity as indicated by BLUPs, compared to six species determined to be susceptible with least squares means estimates. The 29 susceptible species were: 1 from Atripliceae, 4 from Camphorosmeae, 1 from Halopepideae, 2 from Polycnemeae, 6 from Salicornieae, 8 from Salsolae, and 7 from Suaedeae. Most species in the genus Salsola, which are all introduced and weedy, were very susceptible and damaged by CGS. Statistical comparisons and contrasts of BLUPs indicated that these Salsola species were significantly more susceptible than non-target species, including 15 species from relatives in the closely-related genera Bassia (=Kochia), Nitrophila, Salicornia, Sarcocornia, and Suaeda. Of the 29 susceptible species, 10 native or commercially important species in N. America were identified as needing additional tests to determine the extent of any damage caused by infection.     

<Emphasis Type="Italic">mab-31</Emphasis> and the TGF-β pathway act in the ray lineage to pattern <Emphasis Type="Italic">C. elegans</Emphasis>male sensory rays

Background  

C. elegans TGF-β-like Sma/Mab signaling pathway regulates both body size and sensory ray patterning. Most of the components in this pathway were initially identified by genetic screens based on the small body phenotype, and many of these mutants display sensory ray patterning defect. At the cellular level, little is known about how and where these components work although ray structural cell has been implicated as one of the targets. Based on the specific ray patterning abnormality, we aim to identify by RNAi approach additional components that function specifically in the ray lineage to elucidate the regulatory role of TGF-β signaling in ray differentiation.     

Ultrastructural localization of the high molecular weight proteins associated with in vitro-assembled brain microtubules

Microtubules isolated from brain extracts by in vitro assembly (1, 19, 23) are composed principally of two tubulins and two high molecular weight proteins (microtubule-associated proteins [MAPS] 1 and 2) (2,5,7,20). Recently, it was demonstrated that in vitro-assembled brain microtubules (neurotubules) are coated with filaments (5, 7) which are similar to the filaments attached to neurotubules in situ (4, 15, 21, 24, 25), and it was suggested that the filaments are composed of the higher molecular weight MAPs (5, 7, 12). In this study, microtubules were assembled in the presence and absence of the MAPs, and thin sections of the microtubules were examined by electron microscopy. The results show that the filaments only occur on microtubules assembled in the presence of the MAPs and it is therefore concluded that the filaments are composed of the high molecular weight MAP's.     

Supplemental risk evaluations of Puccinia jaceae var. solstitialis for biological control of yellow starthistle

Additional tests of native North American Cirsium species, Saussurea americana, and modern safflower cultivars (Carthamus tinctorius) were requested by regulators and specific interest groups during the risk assessment of foreign isolates of Puccinia jaceae var. solstitialis for biological control of yellow starthistle (YST, Centaurea solstitialis) in the United States. These tests supplement an earlier, extensive host range determination that established P. jaceae from YST as generally host specific and potentially useful for biological control. The additional research was in response to potential hazards identified in an earlier study, changes in safflower cultivars, and concern that P. jaceae might cause a safflower seedling disease similar to hypocotyl infections from infestation by Puccinia carthami teliospores. S. americana, a close relative of yellow starthistle, had not been tested previously. All tests were conducted in a containment greenhouse. Foliage of 19 Cirsium species, 11 safflower cultivars, and S. americana was inoculated with urediniospores and subjected to a 16-h dew period at 18–20 °C. Neither the Cirsium species nor S. americana became infected after foliar inoculations. Compared to foliar infections by P. carthami from safflower in California, only minor infections developed from inoculations with P. jaceae. These were similar to infections observed in earlier studies, and it was not possible to maintain P. jaceae under optimal greenhouse conditions on safflower foliage. Quantitative teliospore inoculations with P. jaceae did not cause infection on safflower hypocotyls, even though large cankers occurred on plants inoculated with P. carthami teliospores. Clear microscopic evidence of infection also was observed in hypocotyls inoculated with P. carthami. These data suggest that native (including rare, threatened, or endangered) Cirsium spp., modern safflower cultivars, and S. americana are not likely to be adversely affected by the use of P. jaceae for biological control of YST. Results from these studies substantiate previous findings and were incorporated in a proposal for permission to use P. jaceae for YST control in California.