Tomasz Płoszaj

mtDNA from the Early Bronze Age to the Roman Period Suggests a Genetic Link between the Indian Subcontinent and Mesopotamian Cradle of Civilization

Ancient DNA methodology was applied to analyse sequences extracted from freshly unearthed remains (teeth) of 4 individuals deeply deposited in slightly alkaline soil of the Tell Ashara (ancient Terqa) and Tell Masaikh (ancient Kar-Assurnasirpal) Syrian archaeological sites, both in the middle Euphrates valley. Dated to the period between 2.5 Kyrs BC and 0.5 Kyrs AD the studied individuals carried mtDNA haplotypes corresponding to the M4b1, M49 and/or M61 haplogroups, which are believed to have arisen in the area of the Indian subcontinent during the Upper Paleolithic and are absent in people living today in Syria. However, they are present in people inhabiting today’s Tibet, Himalayas, India and Pakistan. We anticipate that the analysed remains from Mesopotamia belonged to people with genetic affinity to the Indian subcontinent since the distribution of identified ancient haplotypes indicates solid link with populations from the region of South Asia-Tibet (Trans-Himalaya). They may have been descendants of migrants from much earlier times, spreading the clades of the macrohaplogroup M throughout Eurasia and founding regional Mesopotamian groups like that of Terqa or just merchants moving along trade routes passing near or through the region. None of the successfully identified nuclear alleles turned out to be ΔF508 CFTR, LCT-13910T or Δ32 CCR5.

ARTD1 regulates osteoclastogenesis and bone homeostasis by dampening NF-κB-dependent transcription of IL-1β

While ADP-ribosyltransferase diphtheria toxin-like 1 (ARTD1, formerly PARP1) and its enzymatic activity have been shown to be important for reprogramming and differentiation of cells, such as during adipogenesis, their role and mechanism in regulating osteoclastogenesis and bone homeostasis are largely unknown. Here, in cell culture-based RANKL-induced osteoclastogenesis models, we show that silencing of ARTD1 or inhibition of its enzymatic activity enhances osteoclast differentiation and function. As a consequence of ARTD1 silencing or inhibition, the recruitment of p65/RelA to the IL-1β promoter, which is associated with transcriptionally active histone marks, IL-1β expression and inflammasome-dependent secretion of IL-1β are enhanced. This subsequently promotes sustained induction of the transcription factor Nfatc1/A and osteoclastogenesis in an autocrine manner via the IL-1 receptor. In vivo, Artd1-deficient mice display significantly decreased bone mass as a consequence of increased osteoclast differentiation. Accordingly, the expression of osteoclast markers is enhanced in mutant compared to wild-type mice. Together, these results indicate that ARTD1 controls osteoclast development and bone remodelling via its enzymatic activity by modulating the epigenetic marks surrounding the IL-1β promoter and expression of IL-1β and subsequently also Nfatc1/A.

Between the Baltic and Danubian Worlds: The Genetic Affinities of a Middle Neolithic Population from Central Poland

For a long time, anthropological and genetic research on the Neolithic revolution in Europe was mainly concentrated on the mechanism of agricultural dispersal over different parts of the continent. Recently, attention has shifted towards population processes that occurred after the arrival of the first farmers, transforming the genetically very distinctive early Neolithic Linear Pottery Culture (LBK) and Mesolithic forager populations into present-day Central Europeans. The latest studies indicate that significant changes in this respect took place within the post-Linear Pottery cultures of the Early and Middle Neolithic which were a bridge between the allochthonous LBK and the first indigenous Neolithic culture of north-central Europe—the Funnel Beaker culture (TRB). The paper presents data on mtDNA haplotypes of a Middle Neolithic population dated to 4700/4600–4100/4000 BC belonging to the Brześć Kujawski Group of the Lengyel culture (BKG) from the Kuyavia region in north-central Poland. BKG communities constituted the border of the “Danubian World” in this part of Europe for approx. seven centuries, neighboring foragers of the North European Plain and the southern Baltic basin. MtDNA haplogroups were determined in 11 individuals, and four mtDNA macrohaplogroups were found (H, U5, T, and HV0). The overall haplogroup pattern did not deviate from other post-Linear Pottery populations from central Europe, although a complete lack of N1a and the presence of U5a are noteworthy. Of greatest importance is the observed link between the BKG and the TRB horizon, confirmed by an independent analysis of the craniometric variation of Mesolithic and Neolithic populations inhabiting central Europe. Estimated phylogenetic pattern suggests significant contribution of the post-Linear BKG communities to the origin of the subsequent Middle Neolithic cultures, such as the TRB.

Hunting for the LCT-13910*T Allele between the Middle Neolithic and the Middle Ages Suggests Its Absence in Dairying LBK People Entering the Kuyavia Region in the 8th Millennium BP

Populations from two medieval sites in Central Poland, Stary Brześć Kujawski-4 (SBK-4) and Gruczno, represented high level of lactase persistence (LP) as followed by the LCT-13910*T allele’s presence (0.86 and 0.82, respectively). It was twice as high as in contemporaneous Cedynia (0.4) and Śródka (0.43), both located outside the region, higher than in modern inhabitants of Poland (0.51) and almost as high as in modern Swedish population (0.9). In an attempt to explain the observed differences its frequency changes in time were followed between the Middle Neolithic and the Late Middle Ages in successive dairying populations on a relatively small area (radius ∼60km) containing the two sites. The introduction of the T allele to Kuyavia 7.4 Ka BP by dairying LBK people is not likely, as suggested by the obtained data. It has not been found in any of Neolithic samples dated between 6.3 and 4.5 Ka BP. The identified frequency profile indicates that both the introduction and the beginning of selection could have taken place approx. 4 millennia after first LBK people arrived in the region, shifting the value of LP frequency from 0 to more than 0.8 during less than 130 generations. We hypothesize that the selection process of the T allele was rather rapid, starting just after its introduction into already milking populations and operated via high rates of fertility and mortality on children after weaning through life-threatening conditions, favoring lactose-tolerant individuals. Facing the lack of the T allele in people living on two great European Neolithization routes, the Danubian and Mediterranean ones, and based on its high frequency in northern Iberia, its presence in Scandinavia and estimated occurrence in Central Poland, we propose an alternative Northern Route of its spreading as very likely. None of the successfully identified nuclear alleles turned out to be deltaF508 CFTR.

Frequency of lactase persistence genotype in a healthy Polish population

The majority of mammals are unable to digest lactose due to post-weaning deactivation of the LCT gene, which is responsible for encoding the enzyme lactase (i.e., adult-type hypolactasia). A substitution of C with T at position −13910 bp upstream of the LCT gene has been linked to the lactase persistence phenotype in European populations. We investigated the frequency of LCT-13910C>T polymorphism in 223 blood donors from central Poland. All samples were genotyped by polymerase chain reaction and direct sequencing. The LCT-13910 T allele (lactase persistence) was present in 51% of individuals sampled from the Polish population. We did not find any non-European variants associated with lactase persistence (LCT-13907C>G, LCT-13913T>C, LCT-13915T>G), or any new polymorphisms within the sequenced region. Allele frequencies obtained are in agreement with results from other countries and confirm the unique pattern of distribution of the LCT-13910C>T genotype in Europe.

The Late Neolithic sepulchral and ritual place of site 14 in Kowal (Kuyavia, Central Poland)

Zusammenfassung: Forschungsgegenstand dieses Artikels ist ein Begräbnis- und Ritualplatz der Kugelamphoren-Kultur der Fundstätte 14 in Kowal (Zentral-Polen). Die Stätte umfasst einen Submegalithen sowie einen Ritualplatz mit Tiergräbern und einer menschlichen Bestattung. Der Komplex kann auf die Zeit zwischen 3250/3100 und 2400/2150 v. Chr. datiert werden. Analysen haben gezeigt, dass der Platz über eine lange Zeit benutzt wurde, und dass die durchgeführten Rituale sehr komplex waren. Beweise wurden gefunden, dass an einigen Stellen rituelle Handlungen ohne Unterbrechung über viele Jahre durchgeführt wurden und einzelne Tieropfer und Niederlegungen nur durch einen Zeitraum von wenigen Jahren voneinander getrennt waren. Die Fund-Struktur einiger Gruben zeigt eine sehr penible Organisation des geheiligten Platzes und ermöglicht an einigen Stellen sogar eine Unterscheidung in Nutz- und rituelle Zonen. Weitere wichtige Informationen wurden durch die Untersuchung einer menschlichen Grablege an dieser Stätte gewonnen. Die Analyse der Grabbeigaben hat deren Natur als rituelle Gaben gezeigt; weiter weist sie auf eine dominante Rolle von Tieren im täglichen Leben der verstorbenen Person hin. Wichtige Daten wurden durch die paläo-biologische Untersuchung der menschlichen Knochen gewonnen. Die paläo-serologische Analyse ergab die Blutgruppe der bestatteten Person. Dies ist der erste derartige Nachweis in der Geschichte der europäischen neolithischen Forschung. Bei den Ergebnissen der DNA-Analyse handelt es eines Vertreters der Kugelamphoren-Kultur in der Literatur und zudem um eine von nur wenigen solcher Untersuchungen eines neolithischen Menschen. Einige der nachgewiesenen Allele sind die ältesten Beispiele dieses speziellen Allel-Typs, die in menschlichen Überresten bis heute identifiziert wurden. Die DNA-Analysen belegen eine Laktose-Intoleranz der bestatteten Person, sie legen weiter eine genetische Verwandtschaft mit der Bevölkerung aus der Region des Fruchtbaren Halbmondes nahe. Die Analyse des stabilen Sauerstoff-Isotopen-Verhältnisses zeigt Muster für häufigeren Ortswechsel und das Entwöhnungsalter. Analysen von stabilen Kohlenstoff- und Stickstoff-Isotopen-Verhältnissen haben Rückschlüsse auf die Ernährung ermöglicht; besondere Aufmerksamkeit verdient die Tatsache, dass bis zu 25 % der Kohlenhydrat-haltigen Nahrung aus Hirse bestanden hat. Das ist der früheste Isotopen-gestützte Nachweis von Hirse in der menschlichen Ernährung in Mitteleuropa. Résumé: Le site 14 de Kowal en Pologne centrale, un complexe rituel appartenant à la culture des amphores globulaires, forme le sujet de cet article. Le site comporte un sub-mégalithe, une structure rituelle avec des dépôts d’animaux et une sépulture humaine. Il fut occupé pendant une longue période allant de 3250/3100 à 2400/2150 av. J.-C. et les rituels étaient complexes: certains furent célébrés au même endroit pendant longtemps, tandis que les sacrifices et dépôts d’animaux n’étaient séparés que par de courts intervalles de peutêtre quelques années. Le remplissage de plusieurs fosses démontre une organisation précise de l’espace sacré, et il a même été possible de faire une distinction entre les actes de déposition rituelle et les dépôts de type utilitaire à l’intérieur de ces fosses. L’étude de la sépulture humaine a également fourni d’importants indices. Il en ressort qu’un individu intentionnellement choisi parmi les membres de ‘l’aristocratie’ locale a été la victime d’un meurtre rituel. L’analyse du mobilier funéraire indique qu’il s’agit d’offrandes rituelles et suggère que les animaux jouaient un rôle déterminant dans la vie de cet individu. L’étude de ses ossements a fourni un important ensemble de données, y compris les résultats d’une analyse sérologique permettant d’identifier – pour la première fois pour le Néolithique européen – le groupe sanguin de cette homme. Dans une étude également pionnière pour la culture des amphores globulaires, l’analyse ADN a permis de décrire la composition génétique d’un de ses membres, une rareté dans les études sur le Néolithique. Certains allèles sont les plus anciens exemplaires d’un type particulier d’allèles jusqu’à présent découvert dans des restes humains. Les données de l’analyse de l’ADN de cet individu suggèrent qu’il était intolérant au lactose et qu’il avait des affinités génétiques avec le Croissant fertile. Les résultats de l’analyse des isotopes stables de l’oxygène peuvent être interprétés soit en termes de mobilité ou de sevrage. Ceux provenant de l’analyse des isotopes stables du carbone et de l’azote nous permettent de reconstruire l’alimentation de notre individu ; en particulier le millet fournissait jusqu’à 25 % de sa consommation en glucides. Ce résultat constitue le premier indice isotopique de la présence du millet dans l’alimentation des populations de l’Europe centrale. Abstract: Site 14 in Kowal (central Poland), a funerary and ritual place of the Globular amphora culture forms the subject of this article. The site includes a sub-megalith, a ritual feature with animal burials and a human burial. The complex can be dated to a period spanning from 3250/3100 to 2400/2150 BC; it was used over a long period, and the rituals carried out were complex. Rituals were performed over many years in certain areas, while individual acts of animal sacrifice and deposition may have been separated by a short interval of just a few years. The fill of some pits shows that the sacred space was carefully organised and it has been possible to distinguish between some utilitarian and ritual deposits within them. Other important information is provided by the study of a human burial discovered at the site. It is possible that a deliberately chosen individual from the tribal ‘aristocracy’ was the victim of a ritual murder. The analysis of the grave goods reveals that they were ritual offerings, and these suggest that animals played a dominant role in the deceased man’s daily life. The study of his bones provides important new insights, including those from a palaeo-serological analysis revealing his blood group, a first in the history of European Neolithic research. The results of DNA analysis constitute the first description of the genetic traits of a representative of the Globular amphora culture, among very few such DNA profiles available for a Neolithic person. Some of the alleles discovered are the oldest examples of particular alleles among those identified in human remains so far. DNA evidence suggests that the individual was lactose intolerant, and it demonstrates a genetic relationship with the Fertile Crescent. The analysis of stable oxygen isotope ratios shows patterns compatible with either residential mobility or weaning age. The analysis of stable carbon and nitrogen isotope ratios allowed us to reconstruct the individual’s diet; particular attention is drawn to the fact that up to 25 % of his carbohydrate diet consisted of millet. This is the earliest isotopic evidence for the presence of millet in the diet of people living in Central Europe.

Origin of the ornamented bâton percé from the Gołębiewo site 47 as a trigger of discussion on long-distance exchange among Early Mesolithic communities of Central Poland and Northern Europe

This article describes evidence for contact and exchange among Mesolithic communities in Poland and Scandinavia, based on the interdisciplinary analysis of an ornamented bâton percé from Gołębiewo site 47 (Central Poland). Typological and chronological-cultural analyses show the artefact to be most likely produced in the North European Plain, during the Boreal period. Carbon-14 dating confirms the antiquity of the artefact. Ancient DNA analysis shows the artefact to be of Rangifer tarandus antler. Following this species designation, a dispersion analysis of Early-Holocene reindeer remains in Europe was conducted, showing this species to exist only in northern Scandinavia and north-western Russia in this period. Therefore, the bâton from Gołębiewo constitutes the youngest reindeer remains in the European Plain and south-western Scandinavia known to date. An attempt was made to determine the biogeographic region from which the antler used to produce the artefact originates from. To this end, comprehensive δ18O, δ13C and δ15N isotope analyses were performed. North Karelia and South Lapland were determined as the most probable regions in terms of isotopic data, results which correspond to the known distribution range of Rangifer tarandus at this time. In light of these finds, the likelihood of contact between Scandinavia and Central Europe in Early Holocene is evaluated. The bâton percé from Gołębiewo is likely key evidence for long-distance exchange during the Boreal period.

PARP1 promoter links cell cycle progression with adaptation to oxidative environment

Although electrophiles are considered as detrimental to cells, accumulating recent evidence indicates that proliferating non-cancerous and particularly cancerous cells utilize these agents for pro-survival and cell cycle promoting signaling. Hence, the redox shift to mild oxidant release must be balanced by multiple defense mechanisms. Our latest findings demonstrate that cell cycle progression, which dictates oxidant level in stress-free conditions, determines PARP1 transcription. Growth modulating factors regulate CDK4/6-RBs-E2Fs axis. In cells arrested in G1 and G0, RB1-E2F1 and RBL2-E2F4 dimers recruit chromatin remodelers such as HDAC1, SWI/SNF and PRC2 to condense chromatin and turn off transcription. Release of retinoblastoma-based repressive complexes from E2F-dependent gene promoters in response to cell transition to S phase enables transcription of PARP1. This enzyme contributes to repair of oxidative DNA damage by supporting several strand break repair pathways and nucleotide or base excision repair pathways, as well as acting as a co-activator of transcription factors such as NRF2 and HIF1a, which control expression of antioxidant enzymes involved in removal of electrophiles and secondary metabolites. Furthermore, PARP1 is indispensible for transcription of the pro-survival kinases MAP2K6, ERK1/2 and AKT1, and for maintaining MAPK activity by suppressing transcription of the MAPK inhibitor, MPK1. In summary, cell cycle controlled PARP1 transcription helps cells to adapt to a pro-oxidant redox shift.

Searching for signals of recent natural selection in genes of the innate immune response - ancient DNA study

The last decade has seen sharp progress in the field of human evolutionary genetics and a great amount of genetic evidence of natural selection has been provided so far. Since host-pathogen co-evolution is difficult to trace due to the polygenic nature of human susceptibility to microbial diseases, of particular interest is any signal of natural selection in response to the strong selective pressure exerted by pathogens. Analysis of ancient DNA allows for the direct insight into changes of a gene pool content over time and enables monitoring allele frequency fluctuations. Among pathogenic agents, mycobacteria are proved to have remained in an intimate, long-lasting relation with humans, reflected by the current high level of host resistance. Therefore, we aimed to investigate the prevalence of several polymorphisms within innate immune response genes related to susceptibility to mycobacterial diseases (in SLC11A1, MBL2, TLR2, P2RX7, IL10, TNFA) in time series data from North and East Poland (1st-18th century AD, n = 207). The comparison of allele frequencies over time revealed a predominant role of genetic drift in shaping past gene pool of small, probably isolated groups, which was explained by the high level of population differentiation and limited gene flow. However, the trajectory of frequency fluctuations of two SNPs suggested the possibility of their non-neutral evolution and the results of applied forward simulations further strengthened the hypothesis of natural selection acting on those loci. However, we observed an unusual excess of homozygosity in the profile of several SNPs, which pinpoints to the necessity of further research on temporally and spatially diverse samples to support our inference on non-stochastic evolution, ideally employing pathway-based approaches. Nevertheless, our study confirms that time series data could help to decipher very recent human adaptation to life-threatening pathogens and assisting demographic events.

Presence of archaea and selected bacteria in infected root canal systems

Infections of the root canal have polymicrobial etiology. The main group of microflora in the infected pulp is bacteria. There is limited data that archaea may be present in infected pulp tissue. The aim of this study was to check the prevalence of archaea in necrotic root canal samples obtained from patients with primary or post-treatment infection. The prevalence of selected bacteria species (Prevotella intermedia, Porphyromonas gingivalis, Tannerella forsythia, Treponema denticola, Synergistes sp.) in necrotic samples was evaluated as well. Sixty-four samples from root canal were collected for DNA and RNA extraction. A PCR assay based on the 16S rRNA gene was used to determine the presence of archaea and selected bacteria. Of the 64 samples, 6 were analyzed by semiquantitative reverse transcription PCR to estimate expression profiles of 16S rRNA, and another 9 were selected for direct sequencing. Archaea were detected in 48.4% samples. Statistical analysis indicated a negative association in coexistence between archaea and Treponema denticola (P < 0.05; Pearsons χ2 test). The main representative of the Archaea domain found in infected pulp tissue was Methanobrevibacter oralis. Archaea 16S rRNA gene expression was significantly lower than Synergistes sp., Porphyromonas gingivalis, and Tannerella forsythia (P < 0.05; Students t test). Thus, it can be hypothesized that archaea may participate in the endodontic microbial community.