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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 (LCORL) retrocopies were present in all equids but absent from other extant perissodactyls. The majority of LCORL transcripts in horses and donkeys originate from the retrocopies. The initial LCORL 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 LCORL retrocopy segmental amplification in the Equidae family, high expression levels and the ancient timeline for LCORL retrotransposition support a functional role for this structural variant.

A 10-year-old intact female Chinese Crested dog was presented for evaluation and further diagnostics due to persistent symptoms of vulvar swelling, vaginal discharge, and an 8-year history of acyclicity. At presentation, generalized hyperpigmentation and truncal alopecia were identified, with no aberrations of the female phenotype. Vaginal cytology confirmed the influence of estrogen at multiple veterinary visits, and hormonal screening of progesterone and anti-Mullerian hormone indicated gonadal presence. Based on findings from abdominal laparotomy and gonadectomy, the tissue was submitted for histopathology. Histopathologic evaluation identified the gonads to be abnormal testes containing multiple Sertoli and interstitial (Leydig) cell tumors. The histopathologic diagnosis of testes and concurrent normal external female phenotype in the patient lead to a diagnosis of a disorder of sexual development (DSD). Karyotype evaluation by conventional and molecular analysis revealed a two cell line chimeric pattern of 78,XX (80%) and 78,XY (20%) among blood leukocytes, as well as a positive PCR test for the Y-linked SRY gene. Cytogenetic analysis of skin fibroblasts revealed the presence of 78,XX cells exclusively, and PCR tests for the Y-linked SRY gene were negative in the hair and skin samples. These results are consistent with an XX/XY blood chimerism. This is one of the few case reports of a canine with the diagnosis of leukocyte chimerism with normal female phenotypic external genitalia. This case illustrates a distinct presentation for hormonally active Sertoli cell tumorigenesis and demonstrates surgery as a curative treatment option for clinically affected patients.

Structural rearrangements like copy number variations in the male-specific Y chromosome have been associated with male fertility phenotypes in human and mouse but have been sparsely studied in other mammalian species. Here, we designed digital droplet PCR assays for 7 horse male-specific Y chromosome multicopy genes and SRY and evaluated their absolute copy numbers in 209 normal male horses of 22 breeds, 73 XY horses with disorders of sex development and/or infertility, 5 Przewalski's horses and 2 kulans. This established baseline copy number for these genes in horses. The TSPY gene showed the highest copy number and was the most copy number variable between individuals and breeds. SRY was a single-copy gene in most horses but had 2-3 copies in some indigenous breeds. Since SRY is flanked by 2 copies of RBMY, their copy number variations were interrelated and may lead to SRY-negative XY disorders of sex development. The Przewalski's horse and kulan had 1 copy of SRY and RBMY. TSPY and ETSTY2 showed significant copy number variations between cryptorchid and normal males (P

Supplemental Material, Combined_supplemental_materials-Muscatello_et_al for HER2 Overexpression and Amplification in Feline Pulmonary Carcinoma by Luisa Vera Muscatello, Enrico Di Oto, Miriam Dignazzi, William J. Murphy, Ilaria Porcellato, Raffaella De Maria, Terje Raudsepp, Maria Pia Foschini, Monica Sforna, Cinzia Benazzi and Barbara Brunetti in Veterinary Pathology

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Dynamic evolutionary processes and complex structure make the Y chromosome among the most diverse and least understood regions in mammalian genomes. Here, we present an annotated assembly of the male specific region of the horse Y chromosome (eMSY), representing the first comprehensive Y assembly in odd-toed ungulates. The eMSY comprises single-copy, equine specific multi-copy, PAR transposed, and novel ampliconic sequence classes. The eMSY gene density approaches that of autosomes with the highest number of retained X–Y gametologs recorded in eutherians, in addition to novel Y-born and transposed genes. Horse, donkey and mule testis RNAseq reveals several candidate genes for stallion fertility. A novel testis-expressed XY ampliconic sequence class, ETSTY7, is shared with the parasite Parascaris genome, providing evidence for eukaryotic horizontal transfer and inter-chromosomal mobility. Our study highlights the dynamic nature of the Y and provides a reference sequence for improved understanding of equine male development and fertility., The rapidly evolving Y chromosome accumulates male-benefit genes but is often poorly characterized in many mammals. Here, the authors assemble the male specific region of the horse Y chromosome and investigate its evolution and function.

Figure S1. Equine keratinocytes (EK-100) were cultured in various culture conditions. Representative phase contrast images of primary equine keratinocytes (EK-100) cultured in (a) co-culture with irradiated fibroblasts+ 10 uM Y-27632, (b) F + 10 uM Y-27632 (c,d) CNT with or without 10 uM Y-27632 and (e)KSFM+ 10 uM. All images were taken on and day7 following initial culture without passage (×10 magnification. Size bars = 400 μm). Top right images show enlarged magnification (×40 magnification, size bars = 100 μm). Figure S2. Fluorescence-activated cell sorting (FACS) analysis of human keratinocytes (HFK) and mouse fibroblasts (j2) using pan-cytokeratin antibody. HFK cells were incubated without (a) pan-CK antibody or (b) with pan-CK antibody, (c) J2 fibroblasts without pan-CK antibody, or (d) with pan-CK antibody. Figure S3. Validation of antibodies for equine tissues. Specificity and reactivity of CK-14 was tested by using diluted concentration of CK14 1:600, 1:5000, and no antibody respectively in (a) breast cancer tissue and (b) equine skin tissue. All images (×40 magnification, scale bar = 100 μm) are representative of three experimental repeats. (PPTX 5023 kb)

Väitekirja elektrooniline versioon ei sisalda publikatsioone., Elektrit juhtivad polümeerid on eriliste omadustea materjalide klass, mille populaarsus on kasvanud viimase paarikümne aasta jooksul – eelkõige tänu nende valmistamise lihtsusele ning perspektiivsetele tehnoloogilistele rakendustele. On hästi teada, et polümeerid (plastmassid) ei juhi elektrit. Avastus, et mõned polümeerid võivad olla pooljuht omadustega või isegi juhid (juhtivad omadused võrreldavad metallidega) oli algtõukeks uuele teadusvaldkonna aengule – juhtivate polümeeride uurimisele. Nimetatud valdkonna avastamist ja arendamist on maailmas peetud niivõrd oluliseks, et 2000 aasta Nobeli preemia anti just juhtivate plümeeride uurijatele: Alan. J. Heegerile, Alan. G. MacDiarmid,ile ning Hideki Shirakawale polüatsetüleeni avastamise eest 1977 aastal. Juhtivate polümeeride peamised köitvad eelised võrreldes teiste juhtivate materjalidega on nende potentsiaalne töötlemise lihtsus, suhteliselt hea mehaaniline vastupidavus, kõrge elektrijuhtivus, struktuuri paindlikkus, kergus ning praeguse hetkeni juba teadaolevad atraktiivsed ja huvitavd tehnoloogilised rakendused. Juhtivatest polümeeridest üks kõige lootustandvamaid on polüpürrool, tänu oma püsivale ning heale elektrijutivusele ning stabiilsusele võrreldes teiste juhtivate polümeeridega. Lisaks on polüpürrool osutunud bioühilduvaks, mistõttu on teda võimalik kasutada proteesides, kunstlihastes ning närvikudede sidumiseks. Juhtivate polümeeride võimalikke rakendusi on palju ning nende arv pidevalt kasvab, kuna antud materjalide uurimine ning omaduste täiendamine jätkub. Juhtivaid polümeere on kasutatud väga erinevates valdkondades näiteks meditsiinis, sõjaväes, pooljuhttoodete tööstuses. Märkimisväärseid jõupingutusi on tehtud nende rakendamiseks sensorites, implantaatides, polümeersetes patareides, keemilistes vooluallikates. Veel on kasutatud juhtivaid polümeere korrosiooni kaitses, sõjaväes kaitseriietuste valmistamisel, antistaatiliste kattematerjalide valmistamisel ning ioone selekteerivate ja lahutavate membraanide valmistamisel jne. Elektrokeemilise sünteesi käigus sisenevad polümeeri anioonid (dopandid), mis peale langu tasakaalustamise rolli omavad suurt mõju tekkiva polümeeri struktuurile ja sellega omakorda teistele omadustele. Käesolevas töös uuriti detailselt kolme aromaatse-aniooni: benseensulfonaat-aniooniga (BS); para-tolueensulfonaat-aniooniga ja naftaleen-1,5-disulfonaat-aniooniga dopeeritud polüpürrooli (PPy) elektrokeemilisi omadusi vesilahustes. Erineva paksusega kilesid sünteesiti elektrokeemiliselt konstantsel voolutihedusel. Põhiliseks kasutatavaks uurimismeetodiks oli tsükliline voltamperomeetria. Täiendavat informatsiooni saadi mikroanalüüsist (EDS), kvantkeemilistest arvutustest ning pinnauuringutest aatomjõu mikroskoopia meetodiga (AFM.). Eksperimentaalsed uuringud näitasid, et need võrdlemisi sarnaste dopantioonidega (benseensulfonaat vs para-tolueensulfonaat) sünteesitud kiled omavad üsna erinevaid elektrokeemilisi omadusi sõltuvalt lahuse koostisest ning paksusest. Elektrokeemilised mõõtmised näitasid, et aromaatsete dopantidega dopeeritud kilede redoksaktiivsust on võimalik oluliselt tõsta ümberdopeerimisega 2 korda ning suurim redoksaktiivsus saavutati kõikide kilede korral naatriumsulfaati sisaldavas lahuses. Kahjuks polümeeride korral ei realiseeru kogu laeng s.t kogu polümeer ei ole elektrokeemiliselt aktiivne. Ümberdopeerimise tulemused näitasid aga seda, et on võimalik saavutada olukord, kus kogu polümeer on elektrokeemiliselt aktiivne, mitõttu on antud kiled perspektiivsed kasutada keemilistes voolualliktes. Mõõtmistulemused näitasid, et dopeerimisastmed antud kilede korral olid suhteliselt kõrged (≈ 0.4) ületades kirjanduses väljapakutud tulemusi., The essential results of the present electrocehmical study of polypyrrole films doped originally with aromatic anions such as BS; pTS and NDS and measured in various types of electrolytes can be listed as follows. • The comparative study of the properties of PPy films doped with three different aromatic sulfonate anions using four methods revealed that these similar-looking anions have quite different behavior as dopants. • It was established, that the process of reoxidation of the PPy film after reduction in solution containing the same anion as used in the electrodeposition solution is slow and the restoration of the initial redox charge needs additional polarization for more than 10 min. Due to the compact structure of PPy films doped with aromatic anions the mobility of anions and cations is restricted which causes irreversibility of redoxprocesses. • The voltammetric measurements show that redoxactivity (in solution containing the same anion as used in the electrodeposition solution) of the polypyrrole films doped with these sulfonate ions increases in the sequence pTS< BS< NDS, mostly due to the different mobility of cations. The mobility of the ions in these films is mainly determined how densely the films are packed, which in turn partly depends on the interactions between the dopant-anion and polymer chains. • The results of voltammetric measurements and XDS analyses show that ion exchange between PPy films and solution is relatively quick. It was established that small inorganic anions as NO3-, CI-, ClO4- can relatively easily replace aromatic sulfonates in PPy/BS and PPy/pTS films simply during 10 min polarization at 0.4 V. Only one potentiodynamic cycle needed for essential increase of the rate and fullness of the self-exchange of the anions. • The highest redoxactivity and doping levels in the case of all films orginally doped with aromatic anions were obtained in solution Na2SO4. The self- exchange ability of the double charged SO42- anion is more pronounced and the exchange process does not need electrochemical activation. • The redoxcapacity and corresponding electrochemical doping level of the redoped PPy films are essentially higher than that before ion exchange, showing that the PPy matrix formed in the presence of sulfonate anions does not collapse after ion exchange and it creates good conditions for the mobility of smaller inorganic anion. • The results of electrochemical measurements show clearly that the PPy films synthesized in the presence of BS, pTS and NDS and redoped by self-exchange with chloride, nitrate, perchlorate and sulfate anions have good redoxcapacity and stability.

Comparative genome analysis is one of the ways to investigate the equine genome. In the present work this was carried out by mapping specific genes to horse chromosomes and by detecting homologous chromosomal segments between horse and other species. Zoo-FISH on equine metaphase chromosomes using human whole chromosome painting probes (WCPs) for the first time delineated homologous segments between the karyotypes of the two species. The homology was further refined using microdissected chromosome arm specific paints (ASPs) for HSA2, 5, 6, 16 and 19. The Zoo-FISH studies serve as a basis to extrapolate and efficiently transfer gene mapping data from the advanced gene maps of human/mouse to that of the horse.An important part of any genome mapping project is mapping ofspecific genes. In this thesis four genes were added to the horse physical map: IGF2 using an equine specific probe, and MC1R, KIT, PDGFRA using heterologous (porcine) probes. Successful use of heterologous large insert genomic BAC clones for the assignment of individual genes across distantly related species was the first such report in farm animals, and opens new avenues for developing comparative maps in the absence of species specific gene probes. The comparative gene mapping work was further extended to donkey - another equid species. These results represent the first mapping data in the donkey and provide interesting comparative information with the closely related horse genome.Comparison between the karyotypes of the two equid species was expanded by generating fifteen microdissected equine chromosome specific probes (all meta- and submetacentric autosomes and the sex chromosomes) and hybridizing them to donkey metaphase chromosomes. Eight of the equine WCPs showed one-to-one correspondence with the donkey chromosomes, one to a single arm, while six to 2-3 arms on separate chromosomes. The results provided an insight into how the two genomes are organized in relation to each other, with respect to the equine chromosomes used in the present study. An important outcome of the results was indirect deduction of homology between human and donkey karyotypes. The latter gives a basis for comparison of the donkey genome with other mammalian species.​​​​​​​Accumulating Zoo-FISH data between human and several non-primate species help to understand how different mammalian genomes are organized in relation to each other. Analysis of comparative chromosome painting data between distantly related species enabled identification of evolutionarily conserved whole chromosomes, large chromosomal segments and contiguous synteny combinations, that very likely comprise the karyotype of an eutherian ancestor. The equine genome comprises the focus for all investigations within the thesis, and therefore the results are discussed with special emphasis on the horse.

The unique evolutionary dynamics and complex structure make the Y chromosome the most diverse and least understood region in the mammalian genome, despite its undisputable role in sex determination, development, and male fertility. Here we present the first contig-level annotated draft assembly for the alpaca (Vicugna pacos) Y chromosome based on hybrid assembly of short- and long-read sequence data of flow-sorted Y. The latter was also used for cDNA selection providing Y-enriched testis transcriptome for annotation. The final assembly of 8.22 Mb comprised 4.5 Mb of male specific Y (MSY) and 3.7 Mb of the pseudoautosomal region. In MSY, we annotated 15 X-degenerate genes and two novel transcripts, but no transposed sequences. Two MSY genes, HSFY and RBMY, are multicopy. The pseudoautosomal boundary is located between SHROOM2 and HSFY. Comparative analysis shows that the small and cytogenetically distinct alpaca Y shares most of MSY sequences with the larger dromedary and Bactrian camel Y chromosomes. Most of alpaca X-degenerate genes are also shared with other mammalian MSYs, though WWC3Y is Y-specific only in alpaca/camels and the horse. The partial alpaca Y assembly is a starting point for further expansion and will have applications in the study of camelid populations and male biology.

Elektrit juhtivad polümeerid (edaspidi juhtivad polümeerid) on lihtsasti valmistatavad, nad on suhteliselt vastupidavad, kerged, suure mahtuvuse ja elektrijuhtivusega, mistõttu on nende kasutamine tänasel päeval laialdaselt levinud. Lisaks sellele võivad olla juhtivad polümeerid ka kokkuvõttes odavamad kui mõned muud sarnaste omadustega materjalid. Bakalaureusetöö eesmärk oli uurida sünteesil kasutatava voolutiheduse ja kile paksuse mõju PPy/pTS polümeerkilede elektrokeemiliste omadustele naatriumperkloraati (NaClO4) sisaldavas vesilahuses. Kile elektrokeemilisi omadusi uuriti tsüklilise voltamperomeetria (CV) meetodit kasutades.

Juhtivad polümeerid on kergesti valmistatavad ja töödeldavad, neil on suhteliselt hea mehaaniline vastupidavus, nad on kerged, paindlikud ja suhteliselt kõrge elektrijuhtivusega. Varieerides sünteesitingimusi ning polümeeri lähteained on võimalik luua väga erinevate, eriliste omadustega ja rakendustega polümeermaterjale, mille hinnaklass on sageli odavam kui sarnaste omadustega materjalil. Antud bakalaureusetöö eesmärgiks oli uurida PPy/BzSO3 polümeerkile elektrokeemiliste omaduste sõltuvust mõõtelahuse kontsentratsioonist ning hinnata erinevate ioonide rolli polümeeri oksüdeerimisel ja redutseerimisel.