PLOS ONE: [sortOrder=DATE_NEWEST_FIRST, sort=Date, newest first, q=subject:"Cytogenetic techniques"]PLOShttps://journals.plos.org/plosone/webmaster@plos.orgaccelerating the publication of peer-reviewed sciencehttps://journals.plos.org/plosone/search/feed/atom?sortOrder=DATE_NEWEST_FIRST&sort=Date,+newest+first&unformattedQuery=subject:%22Cytogenetic+techniques%22All PLOS articles are Open Access.https://journals.plos.org/plosone/resource/img/favicon.icohttps://journals.plos.org/plosone/resource/img/favicon.ico2024-03-29T11:24:30ZScreening of the Pandemic Response Box identifies anti-microsporidia compoundsQingyuan HuangJie ChenGuoqing PanAaron W. Reinke10.1371/journal.pntd.00118062023-12-08T14:00:00Z2023-12-08T14:00:00Z<p>by Qingyuan Huang, Jie Chen, Guoqing Pan, Aaron W. Reinke</p>
Microsporidia are fungal obligate intracellular pathogens, which infect most animals and cause microsporidiosis. Despite the serious threat that microsporidia pose to humans and agricultural animals, few drugs are available for the treatment and control of microsporidia. To identify novel inhibitors, we took advantage of the model organism <i>Caenorhabditis elegans</i> infected with its natural microsporidian <i>Nematocida parisii</i>. We used this system to screen the Pandemic Response Box, a collection of 400 diverse compounds with known antimicrobial activity. After testing these compounds in a 96-well format at high (100 μM) and low (40 μM) concentrations, we identified four inhibitors that restored the ability of <i>C</i>. <i>elegans</i> to produce progeny in the presence of <i>N</i>. <i>parisii</i>. All four compounds reduced the pathogen load of both <i>N</i>. <i>parisii</i> and <i>Pancytospora epiphaga</i>, a <i>C</i>. <i>elegans</i>-infecting microsporidia related to human-infecting species. One of these compounds, a known inhibitor of a viral protease, MMV1006203, inhibited invasion and prevented the firing of spores. A bis-indole derivative, MMV1593539, decreased spore viability. An albendazole analog, MMV1782387, inhibited proliferation of <i>N</i>. <i>parisii</i>. We tested albendazole as well as 5 other analogs and observed that MMV1782387 was amongst the strongest inhibitors of <i>N</i>. <i>parisii</i> and displayed the least host toxicity. Our study further demonstrates the effectiveness of the <i>C</i>. <i>elegans-N</i>. <i>parisii</i> system for discovering microsporidia inhibitors and the compounds we identified provide potential scaffolds for anti-microsporidia drug development.Comparative chromosome painting in three Pelecaniformes species (Aves): Exploring the role of macro and microchromosome fusions in karyotypic evolutionIgor Chamon Assumpção SeligmannIvanete de Oliveira FuroMichelly da Silva dos SantosRicardo José GunskiAnalía del Valle GarneroFabio Augusto Oliveira SilvaPatricia O´BrienMalcolm Ferguson-SmithRafael KretschmerEdivaldo Herculano C. de Oliveira10.1371/journal.pone.02947762023-11-27T14:00:00Z2023-11-27T14:00:00Z<p>by Igor Chamon Assumpção Seligmann, Ivanete de Oliveira Furo, Michelly da Silva dos Santos, Ricardo José Gunski, Analía del Valle Garnero, Fabio Augusto Oliveira Silva, Patricia O´Brien, Malcolm Ferguson-Smith, Rafael Kretschmer, Edivaldo Herculano C. de Oliveira</p>
Pelecaniformes is an order of waterbirds that exhibit diverse and distinct morphologies. Ibis, heron, pelican, hammerkop, and shoebill are included within the order. Despite their fascinating features, the phylogenetic relationships among the families within Pelecaniformes remain uncertain and pose challenges due to their complex evolutionary history. Their karyotypic evolution is another little-known aspect. Therefore, to shed light on the chromosomal rearrangements that have occurred during the evolution of Pelecaniformes, we have used whole macrochromosome probes from <i>Gallus gallus</i> (GGA) to show homologies on three species with different diploid numbers, namely <i>Cochlearius cochlearius</i> (2n = 74), <i>Eudocimus ruber</i> (2n = 66), and <i>Syrigma sibilatrix</i> (2n = 62). A fusion between GGA6 and GGA7 was found in <i>C</i>. <i>cochlearius</i> and <i>S</i>. <i>sibilatrix</i>. In <i>S</i>. <i>sibilatrix</i> the GGA8, GGA9 and GGA10 hybridized to the long arms of biarmed macrochromosomes, indicating fusions with microchromosomes. In <i>E</i>. <i>ruber</i> the GGA7 and GGA8 hybridized to the same chromosome pair. After comparing our painting results with previously published data, we show that distinct chromosomal rearrangements have occurred in different Pelecaniformes lineages. Our study provides new insight into the evolutionary history of Pelecaniformes and the chromosomal changes involving their macrochromosomes and microchromosomes that have taken place in different species within this order.Dorsal raphe stimulation relays a reward signal to the ventral tegmental area via GluN2C NMDA receptorsGiovanni HernandezWillemieke M. KouwenhovenEmmanuelle PoirierKarim LebiedDaniel LévesquePierre-Paul Rompré10.1371/journal.pone.02935642023-11-06T14:00:00Z2023-11-06T14:00:00Z<p>by Giovanni Hernandez, Willemieke M. Kouwenhoven, Emmanuelle Poirier, Karim Lebied, Daniel Lévesque, Pierre-Paul Rompré</p>
Background <p>Glutamate relays a reward signal from the dorsal raphe (DR) to the ventral tegmental area (VTA). However, the role of the different subtypes of N-methyl-D-aspartate (NMDA) receptors is complex and not clearly understood. Therefore, we measured NMDA receptors subunits expression in limbic brain areas. In addition, we studied the effects of VTA down-regulation of GluN2C NMDA receptor on the reward signal that arises from DR electrical stimulation.</p> Methods <p>Using qPCR, we identified the relative composition of the different Grin2a-d subunits of the NMDA receptors in several brain areas. Then, we used fluorescent <i>in situ</i> hybridization (FISH) to evaluate the colocalization of Grin2c and tyrosine hydroxylase (TH) mRNA in VTA neurons. To assess the role of GluN2C in brain stimulation reward, we downregulated this receptor using small interfering RNA (siRNA) in rats self-stimulating for electrical pulses delivered to the DR. To delineate further the specific role of GluN2C in relaying the reward signal, we pharmacologically altered the function of VTA NMDA receptors by bilaterally microinjecting the NMDA receptor antagonist PPPA.</p> Results <p>We identified GluN2C as the most abundant subunit of the NMDA receptor expressed in the VTA. FISH revealed that about 50% of TH-positive neurons colocalize with Grin2c transcript. siRNA manipulation produced a selective down-regulation of the GluN2C protein subunit and a significant reduction in brain stimulation reward. Interestingly, PPPA enhanced brain stimulation reward, but only in rats that received the nonactive RNA sequence.</p> Conclusion <p>The present results suggest that VTA glutamate neurotransmission relays a reward signal initiated by DR stimulation by acting on GluN2C NMDA receptors.</p>Genetic diversity analysis in the Brazilian Amazon reveals a new evolutionary lineage and new karyotype for the genus <i>Mesomys</i> (Rodentia, Echimyidae, Eumysopinae)Leony Dias de OliveiraWillam Oliveira da SilvaMarlyson Jeremias Rodrigues da CostaJeferson Costa CarneiroIracilda SampaioJuliane Saldanha da SilvaRogério Vieira RossiAna Cristina Mendes-OliveiraJulio Cesar PieczarkaCleusa Yoshiko Nagamachi10.1371/journal.pone.02917972023-10-04T14:00:00Z2023-10-04T14:00:00Z<p>by Leony Dias de Oliveira, Willam Oliveira da Silva, Marlyson Jeremias Rodrigues da Costa, Jeferson Costa Carneiro, Iracilda Sampaio, Juliane Saldanha da Silva, Rogério Vieira Rossi, Ana Cristina Mendes-Oliveira, Julio Cesar Pieczarka, Cleusa Yoshiko Nagamachi</p>
Morphological, molecular and chromosomal studies in the genera <i>Lonchothrix</i> and <i>Mesomys</i> have contributed to a better understanding of taxonomic design, phylogenetic relationships and karyotypic patterns. Recent molecular investigations have shown a yet undescribed diversity, suggesting that these taxa are even more diverse than previously assumed. Furthermore, some authors have questioned the limits of geographic distribution in the Amazon region for the species <i>M</i>. <i>hispidus</i> and <i>M</i>. <i>stimulax</i>. In this sense, the current study sought to understand the karyotypic evolution and geographic limits of the genus <i>Mesomys</i>, based on classical (G- and C-banding) and molecular cytogenetic analysis (FISH using rDNA 18S and telomeric probes) and through the sequencing of mitochondrial genes Cytochrome b (Cytb) and Cytochrome Oxidase—Subunit I (CO using phylogeny, species delimitation and time of divergence, from samples of different locations in the Brazilian Amazon. The species <i>M</i>. <i>stimulax</i> and <i>Mesomys</i> sp. presented 2n = 60/FN = 110, while <i>M</i>. <i>hispidus</i> presented 2n = 60/FN = 112, hitherto unpublished. Molecular dating showed that <i>Mesomys</i> diversification occurred during the Plio-Pleistocene period, with <i>M</i>. <i>occultus</i> diverging at around 5.1 Ma, followed by <i>Mesomys</i> sp. (4.1 Ma) and, more recently, the separation between <i>M</i>. <i>hispidus</i> and <i>M</i>. <i>stimulax</i> (3.5 Ma). The ABGD and ASAP species delimiters support the formation of 7 and 8 potential species of the genus <i>Mesomys</i>, respectively. Furthermore, in both analyzes <i>Mesomys</i> sp. was recovered as a valid species. Our multidisciplinary approach involving karyotypic, molecular and biogeographic analysis is the first performed in <i>Mesomys</i>, with the description of a new karyotype for <i>M</i>. <i>hispidus</i>, a new independent lineage for the genus and new distribution data for <i>M</i>. <i>hispidus</i> and <i>M</i>. <i>stimulax</i>.Analysis of human invasive cytotrophoblasts demonstrates mosaic aneuploidyJingly F. WeierChristy FerlatteAdolf BaumgartnerHa Nam NguyenBeatrice A. WeierHeinz-Ulrich G. Weier10.1371/journal.pone.02843172023-07-21T14:00:00Z2023-07-21T14:00:00Z<p>by Jingly F. Weier, Christy Ferlatte, Adolf Baumgartner, Ha Nam Nguyen, Beatrice A. Weier, Heinz-Ulrich G. Weier</p>
A total of 24 chromosome-specific fluorescence <i>in situ</i> hybridization probes for interphase nucleus analysis were developed to determine the chromosomal content of individual human invasive cytotrophoblasts derived from <i>in</i> v<i>itro</i> cultured assays. At least 75% of invasive cytotrophoblasts were hyperdiploid and the total number of chromosomes ranged from 47 to 61. The results also demonstrated that these hyperdiploid invasive cytotrophoblasts showed significant heterogeneity. The most copy number gains were observed for chromosomes 13, 14, 15, 19, 21, and 22 with average copy number greater than 2.3. A parallel study using primary invasive cytotrophoblasts also showed a similar trend of copy number changes. Conclusively, 24-chromosome analysis of human non-proliferating cytotrophoblasts (interphase nuclei) was achieved. Hyperdiploidy and chromosomal heterogeneity without endoduplication in invasive cytotrophoblasts may suggest a selective advantage for invasion and short lifespan during normal placental development.<i>PAK1</i> copy number in breast cancer—Associations with proliferation and molecular subtypesAnette H. SkjervoldMarit VallaBorgny YtterhusAnna M. Bofin10.1371/journal.pone.02876082023-06-27T14:00:00Z2023-06-27T14:00:00Z<p>by Anette H. Skjervold, Marit Valla, Borgny Ytterhus, Anna M. Bofin</p>
Introduction <p>P21-activated kinase 1 (<i>PAK1)</i> is known to be overexpressed in several human tumour types, including breast cancer (BC). It is located on chromosome 11 (11q13.5-q14.1) and plays a significant role in proliferation in BC. In this study we aimed to assess <i>PAK1</i> gene copy number (CN) in primary breast tumours and their corresponding lymph node metastases, and associations between <i>PAK1</i> CN and proliferation status, molecular subtype, and prognosis. In addition, we aimed to study associations between CNs of <i>PAK1</i> and <i>CCND1</i>. Both genes are located on the long arm of chromosome 11 (11q13).</p> Methods <p>Fluorescence <i>in situ</i> hybridization for <i>PAK1</i> and Chromosome enumeration probe (CEP)11 were used on tissue microarray sections from a series of 512 BC cases. Copy numbers were estimated by counting the number of fluorescent signals for <i>PAK1</i> and CEP11 in 20 tumour cell nuclei. Pearson’s x<sup>2</sup> test was performed to assess associations between <i>PAK1</i> CN and tumour features, and between <i>PAK1</i> and <i>CCND1</i> CNs. Cumulative risk of death from BC and hazard ratios were estimated in analysis of prognosis.</p> Results <p>We found mean <i>PAK1</i> CN ≥4<6 in 26 (5.1%) tumours, and CN ≥ 6 in 22 (4.3%) tumours. The proportion of cases with copy number increase (mean CN ≥4) was highest among HER2 type and Luminal B (HER2<sup>-</sup>) tumours. We found an association between <i>PAK1</i> CN increase, and high proliferation, and high histological grade, but not prognosis. Of cases with <i>PAK1</i> CN ≥ 6, 30% also had <i>CCND1</i> CN ≥ 6.</p> Conclusions <p><i>PAK1</i> copy number increase is associated with high proliferation and high histological grade, but not with prognosis. <i>PAK1</i> CN increase was most frequent in the HER2 type and Luminal B (HER2<sup>-</sup>) subtype. <i>PAK1</i> CN increase is associated with CN increase of <i>CCND1</i>.</p>Ancient segmentally duplicated <i>LCORL</i> retrocopies in equidsKevin BatcherScarlett VarneyTerje RaudseppMatthew JevitPeter DickinsonVidhya JagannathanTosso LeebDanika Bannasch10.1371/journal.pone.02868612023-06-08T14:00:00Z2023-06-08T14:00:00Z<p>by Kevin Batcher, Scarlett Varney, Terje Raudsepp, Matthew Jevit, Peter Dickinson, Vidhya Jagannathan, Tosso Leeb, Danika Bannasch</p>
LINE-1 is an active transposable element encoding proteins capable of inserting host gene retrocopies, resulting in retro-copy number variants (retroCNVs) between individuals. Here, we performed retroCNV discovery using 86 equids and identified 437 retrocopy insertions. Only 5 retroCNVs were shared between horses and other equids, indicating that the majority of retroCNVs inserted after the species diverged. A large number (17–35 copies) of segmentally duplicated Ligand Dependent Nuclear Receptor Corepressor Like (<i>LCORL</i>) retrocopies were present in all equids but absent from other extant perissodactyls. The majority of <i>LCORL</i> transcripts in horses and donkeys originate from the retrocopies. The initial <i>LCORL</i> retrotransposition occurred 18 million years ago (17–19 95% CI), which is coincident with the increase in body size, reduction in digit number, and changes in dentition that characterized equid evolution. Evolutionary conservation of the <i>LCORL</i> retrocopy segmental amplification in the Equidae family, high expression levels and the ancient timeline for <i>LCORL</i> retrotransposition support a functional role for this structural variant.Aquaporin 5 (AQP5) expression in breast cancer and its clinicopathological characteristicsSe Jin JangChulso Moon10.1371/journal.pone.02707522023-01-27T14:00:00Z2023-01-27T14:00:00Z<p>by Se Jin Jang, Chulso Moon</p>
The role of aquaporin water channels (AQPs) has become an area of great interest in human carcinogenesis. In this report, we have demonstrated the expression of AQP5 in breast cancer by analyzing 591 tissue samples with 7-year follow-ups. By immunochemistry analysis, AQP5 overexpression was observed in 36% (212/591 cases). Then, we have focused on the clinicopathologic variables among cancer tissue samples with strong AQP5 expression (3+ expression, 60/591 cases). The strong AQP5 expression was positively correlated with tumor grade in BCs (p<0.001) and was more frequent in ER/PR-negative BCs than positive ones (14.9% vs. 3.3% and 13.1% vs. 4.8%, respectively, both p<0.001), while Her2/neu-positive status was positively correlated with strong expression of AQP5 (p = 0.005). Of note, breast cancer patients with positive AQP expression (212/591 cases) showed a less favorable breast cancer specific survival rate over 7 years of follow and we further conclude that AQP5 expression is an independent molecular marker associated with worse clinical outcomes. By fluorescence <i>in situ</i> hybridization (FISH), we have identified evidence of gene amplification in 3 of 30 readable breast cancer and further conclude that, in breast cancer, at least some part of AQP5 overexpression is associated with an aberration in the genome level.Novel <i>THPO</i> variant in hereditary thrombocytopenia: A potential candidate variant for predisposition to myeloid neoplasmSeok Ryun KwonMan Jin KimYoung-eun LeeJiwon YunDa-jeong JeongJae Hyeon ParkSunghoon KwonDong Soon Lee10.1371/journal.pone.02716242022-12-19T14:00:00Z2022-12-19T14:00:00Z<p>by Seok Ryun Kwon, Man Jin Kim, Young-eun Lee, Jiwon Yun, Da-jeong Jeong, Jae Hyeon Park, Sunghoon Kwon, Dong Soon Lee</p>
Hereditary thrombocytopenia is a heterogeneous group of congenital disorders with a wide range of symptoms depending on the severity of platelet dysfunction or thrombocytopenia. Because of its clinical phenotypes and the bone marrow morphology associated with this condition, hereditary thrombocytopenia can be misdiagnosed as primary immune thrombocytopenia and myelodysplastic syndrome. Therefore, genetic evidence is necessary for the accurate diagnosis of hereditary thrombocytopenia. Refractory cytopenia of childhood is a subgroup of myelodysplastic syndrome that was added to the World Health Organization classification in 2008. To investigate the germline and somatic variants associated with refractory cytopenia of childhood, we performed targeted multigene sequencing in three patients with refractory cytopenia of childhood. Of the three patients, one progressed from megakaryocytic hypoplasia with thrombocytopenia, and targeted multigene sequencing revealed <i>THPO</i> variants in this patient and his sister. We propose that the monoallelic deletion of <i>THPO</i> is a potential candidate for germline predisposition to myeloid malignancy.Pannexin 1 activity in astroglia sets hippocampal neuronal network patternsFlora VasileElena DossiJulien MoulardPascal EzanLaure LecoinMartine Cohen-SalmonPhilippe MaillyMarc Le BertIsabelle CouillinAlexis BemelmansNathalie Rouach10.1371/journal.pbio.30018912022-12-07T14:00:00Z2022-12-07T14:00:00Z<p>by Flora Vasile, Elena Dossi, Julien Moulard, Pascal Ezan, Laure Lecoin, Martine Cohen-Salmon, Philippe Mailly, Marc Le Bert, Isabelle Couillin, Alexis Bemelmans, Nathalie Rouach</p>
Astroglial release of molecules is thought to actively modulate neuronal activity, but the nature, release pathway, and cellular targets of these neuroactive molecules are still unclear. Pannexin 1, expressed by neurons and astrocytes, form nonselective large pore channels that mediate extracellular exchange of molecules. The functional relevance of these channels has been mostly studied in brain tissues, without considering their specific role in different cell types, or in neurons. Thus, our knowledge of astroglial pannexin 1 regulation and its control of neuronal activity remains very limited, largely due to the lack of tools targeting these channels in a cell-specific way. We here show that astroglial pannexin 1 expression in mice is developmentally regulated and that its activation is activity-dependent. Using astrocyte-specific molecular tools, we found that astroglial-specific pannexin 1 channel activation, in contrast to pannexin 1 activation in all cell types, selectively and negatively regulates hippocampal networks, with their disruption inducing a drastic switch from bursts to paroxysmal activity. This decrease in neuronal excitability occurs via an unconventional astroglial mechanism whereby pannexin 1 channel activity drives purinergic signaling-mediated regulation of hyperpolarisation-activated cyclic nucleotide (HCN)-gated channels. Our findings suggest that astroglial pannexin 1 channel activation serves as a negative feedback mechanism crucial for the inhibition of hippocampal neuronal networks.Co-depletion of NIPBL and WAPL balance cohesin activity to correct gene misexpressionJennifer M. LuppinoAndrew FieldSon C. NguyenDaniel S. ParkParisha P. ShahRichard J. AbdillYemin LanRebecca YunkerRajan JainKaren AdelmanEric F. Joyce10.1371/journal.pgen.10105282022-11-30T14:00:00Z2022-11-30T14:00:00Z<p>by Jennifer M. Luppino, Andrew Field, Son C. Nguyen, Daniel S. Park, Parisha P. Shah, Richard J. Abdill, Yemin Lan, Rebecca Yunker, Rajan Jain, Karen Adelman, Eric F. Joyce</p>
The relationship between cohesin-mediated chromatin looping and gene expression remains unclear. NIPBL and WAPL are two opposing regulators of cohesin activity; depletion of either is associated with changes in both chromatin folding and transcription across a wide range of cell types. However, a direct comparison of their individual and combined effects on gene expression in the same cell type is lacking. We find that NIPBL or WAPL depletion in human HCT116 cells each alter the expression of ~2,000 genes, with only ~30% of the genes shared between the conditions. We find that clusters of differentially expressed genes within the same topologically associated domain (TAD) show coordinated misexpression, suggesting some genomic domains are especially sensitive to both more or less cohesin. Finally, co-depletion of NIPBL and WAPL restores the majority of gene misexpression as compared to either knockdown alone. A similar set of NIPBL-sensitive genes are rescued following CTCF co-depletion. Together, this indicates that altered transcription due to reduced cohesin activity can be functionally offset by removal of either its negative regulator (WAPL) or the physical barriers (CTCF) that restrict loop-extrusion events.Evaluation of a fluorescence <i>in situ</i> hybridization (FISH)-based method for detection of SARS-CoV-2 in salivaGerrit G. TammingaGijsbert J. JansenMarit Wiersma10.1371/journal.pone.02773672022-11-08T14:00:00Z2022-11-08T14:00:00Z<p>by Gerrit G. Tamminga, Gijsbert J. Jansen, Marit Wiersma</p>
The use of a non-invasive fluorescence <i>in situ</i> hybridization (FISH)-based method on saliva for the detection of SARS-CoV-2 is evaluated in a proof-of-concept study and thereafter utilized in an outpatient setting with the Biotrack-MED® analyzer. For a proof-of-concept study, saliva samples were obtained from 28 persons with mild or moderate COVID-19-related symptoms who were tested RT-PCR positive or negative for SARS-CoV-2. In an outpatient setting, 972 individual saliva samples were utilized. All saliva samples were FISHed with a Cy3-labeled SARS-CoV-2-specific DNA probe and were analyzed manually by fluorescence microscopy (proof-of-concept) or with the SARS-CoV-2 application of the Biotrack-MED® analyzer, a semi-autonomous multi-sample filter cytometer. The proof-of-concept study showed a sensitivity of 96.0% and a specificity of 98.5% and is therefore comparable to the RT-PCR analysis of nasopharyngeal swabs. The outpatient setting showed a sensitivity of 90.9% and a specificity of 94.5% and seems therefore a valid assay for the detection of SARS-CoV-2 in individuals that are healthy, mild or moderate symptomatic. In conclusion, the method evaluated in this study, the FISH-based SARS-CoV-2 application of the Biotrack-MED® analyzer, is a sensitive and reliable assay for the detection of SARS-CoV-2 in the general population.Single-cell measurement quality in bitsJayan RammohanSwarnavo SarkarDavid Ross10.1371/journal.pone.02692722022-08-11T14:00:00Z2022-08-11T14:00:00Z<p>by Jayan Rammohan, Swarnavo Sarkar, David Ross</p>
Single-cell measurements have revolutionized our understanding of heterogeneity in cellular response. However, there is no universally comparable way to assess single-cell measurement quality. Here, we show how information theory can be used to assess and compare single-cell measurement quality in bits, which provides a universally comparable metric for information content. We anticipate that the experimental and theoretical approaches we show here will generally enable comparisons of quality between any single-cell measurement methods.A Krüppel-like factor is required for development and regeneration of germline and yolk cells from somatic stem cells in planariansMelanie IssigonisAkshada B. RedkarTania RozarioUmair W. KhanRosa Mejia-SanchezSylvain W. LapanPeter W. ReddienPhillip A. Newmark10.1371/journal.pbio.30014722022-07-15T14:00:00Z2022-07-15T14:00:00Z<p>by Melanie Issigonis, Akshada B. Redkar, Tania Rozario, Umair W. Khan, Rosa Mejia-Sanchez, Sylvain W. Lapan, Peter W. Reddien, Phillip A. Newmark</p>
Sexually reproducing animals segregate their germline from their soma. In addition to gamete-producing gonads, planarian and parasitic flatworm reproduction relies on yolk cell–generating accessory reproductive organs (vitellaria) supporting development of yolkless oocytes. Despite the importance of vitellaria for flatworm reproduction (and parasite transmission), little is known about this unique evolutionary innovation. Here, we examine reproductive system development in the planarian <i>Schmidtea mediterranea</i>, in which pluripotent stem cells generate both somatic and germ cell lineages. We show that a homolog of the pluripotency factor Klf4 is expressed in primordial germ cells (PGCs), presumptive germline stem cells (GSCs), and yolk cell progenitors. Knockdown of this <i>klf4-like</i> (<i>klf4l</i>) gene results in animals that fail to specify or maintain germ cells; surprisingly, they also fail to maintain yolk cells. We find that yolk cells display germ cell–like attributes and that vitellaria are structurally analogous to gonads. In addition to identifying a new proliferative cell population in planarians (yolk cell progenitors) and defining its niche, our work provides evidence supporting the hypothesis that flatworm germ cells and yolk cells share a common evolutionary origin.Circulating tumor cell assay to non-invasively evaluate PD-L1 and other therapeutic targets in multiple cancersRaymond PageDarshana PatilDadasaheb AkolkarSudha S. MurthyKiran BendaleRevati PatilPradeep FulmaliPooja FulmaliArchana AdhavSneha PuranikSachin ApurwaVineet DattaChirantan BoseStefan SchusterJinumary JohnAjay SrinivasanRajan Datar10.1371/journal.pone.02701392022-06-17T14:00:00Z2022-06-17T14:00:00Z<p>by Raymond Page, Darshana Patil, Dadasaheb Akolkar, Sudha S. Murthy, Kiran Bendale, Revati Patil, Pradeep Fulmali, Pooja Fulmali, Archana Adhav, Sneha Puranik, Sachin Apurwa, Vineet Datta, Chirantan Bose, Stefan Schuster, Jinumary John, Ajay Srinivasan, Rajan Datar</p>
Biomarker directed selection of targeted anti-neoplastic agents such as immune checkpoint inhibitors, small molecule inhibitors and monoclonal antibodies form an important aspect of cancer treatment. Immunohistochemistry (IHC) analysis of the tumor tissue is the method of choice to evaluate the presence of these biomarkers. However, a significant barrier to biomarker testing on tissue is the availability of an adequate amount of tissue and need for repetitive sampling due to tumor evolution. Also, tumor tissue testing is not immune to inter- and intra-tumor heterogeneity. We describe the analytical and clinical validation of a Circulating Tumor Cell (CTC) assay to accurately assess the presence of PD-L1 22C3 and PD-L1 28.8, ER, PR and HER2, from patients with solid tumors to guide the choice of suitable targeted therapies. Analytically, the test has high sensitivity, specificity, linearity and precision. Based on a blinded case control study, the clinical sensitivity and specificity for PD-L1 (22C3 and 28.8) was determined to be 90% and 100% respectively. The clinical sensitivity and specificity was 83% and 89% for ER; 80% and 94% for PR; 63% and 89% for HER2 (by ICC); and 100% and 92% for HER2 (by FISH), respectively. The performance characteristics of the test support its suitability and adaptability for routine clinical use.