Cutaneous squamous cell carcinoma (cSCC) is one of the most common and fastest increasing forms of cancer worldwide with metastatic potential. Long noncoding RNAs (lncRNAs) are a group of RNA molecules with essential regulatory functions in both physiological and pathological processes. To investigate the function and mode of action of lncRNA plasmacytoma variant translocation 1 (PVT1) in cSCC. Quantitative reverse transcriptase polymerase chain reaction and single-molecule in situ hybridization were used to quantify the expression level of PVT1 in normal skin, premalignant skin lesions, actinic keratosis (AK) and primary and metastatic cSCCs. The function of PVT1 in cSCC was investigated both in vivo (tumour xenografts) and in vitro (competitive cell growth assay, 5-ethynyl-2'-deoxyuridine incorporation assay, colony formation assay and tumour spheroid formation assay) upon CRISPR-Cas9-mediated knockout of the entire PVT1 locus, the knockout of exon 2 of PVT1, and locked nucleic acid (LNA) gapmer-mediated PVT1 knockdown. RNA sequencing analysis was conducted to identify genes and processes regulated by PVT1. We identified PVT1 as a lncRNA upregulated in cSCC in situ and cSCC, associated with the malignant phenotype of cSCC. We showed that the expression of PVT1 in cSCC was regulated by MYC. Both CRISPR-Cas9 deletion of the entire PVT1 locus and LNA gapmer-mediated knockdown of PVT1 transcript impaired the malignant behaviour of cSCC cells, suggesting that PVT1 is an oncogenic transcript in cSCC. Furthermore, knockout of PVT1 exon 2 inhibited cSCC tumour growth both in vivo and in vitro, demonstrating that exon 2 is a critical element for the oncogenic role of PVT1. Mechanistically, we showed that PVT1 was localized in the cell nucleus and its deletion resulted in cellular senescence, increased cyclin-dependent kinase inhibitor 1 (p21/CDKN1A) expression and cell cycle arrest. Our study revealed a previously unrecognized role for exon 2 of PVT1 in its oncogenic role and that PVT1 suppresses cellular senescence in cSCC. PVT1 may be a potential biomarker and therapeutic target in cSCC.

Protists make up the vast diversity of eukaryotic life and play a critical role in biogeochemical cycling and in food webs. Because of their small size, cryptic life cycles, and large population sizes, our understanding of speciation in these organisms is very limited. We performed population genomic analyses on 153 strains isolated from eight populations of the recently radiated dinoflagellate genus Apocalathium, to explore the drivers and mechanisms of speciation processes. Species of this genus inhabit both freshwater and saline habitats, lakes and seas, and are found in cold temperate environments across the world. RAD sequencing analyses revealed that the populations were overall highly differentiated, but morphological similarity was not congruent with genetic similarity. While geographic isolation was to some extent coupled to genetic distance, this pattern was not consistent. Instead, we found evidence that the environment, specifically salinity, is a major factor in driving ecological speciation in Apocalathium. While saline populations were unique in loci coupled to genes involved in osmoregulation, freshwater populations appear to lack these. Our study highlights that adaptation to freshwater through loss of osmoregulatory genes may be an important speciation mechanism in free-living aquatic protists.

SLE is a systemic autoimmune disease with a large number of common risk gene variants, but several rare gene variants can cause monogenic SLE. The relationship between common and rare variants in SLE is unclear. We therefore investigated the occurrence of rare deleterious variants in patients with childhood-onset SLE (cSLE) and adult-onset SLE (aSLE) and compared the frequency of these variants with their individual SLE polygenic risk score (PRS). Targeted sequencing of 1832 gene regions, including coding regions of 31 genes associated with monogenic SLE, was performed in 958 patients with SLE and 1026 healthy individuals. A total of 116 patients with SLE had disease onset before the age of 18 (cSLE). An SLE common variant PRS was created from 37 SLE genome-wide association study single nucleotide variants (SNVs). Rare coding deleterious SNVs (RD SNVs) were observed in 23 of the monogenic SLE-associated genes. Six per cent of patients with cSLE, compared with 3.2% of controls and 4.6% of patients with aSLE, carried rare deleterious alleles. In cSLE, RD SNVs were observed in the C1S, DDX58, IFIH1, IKZF1, RNASEH2A and C8A genes. A PRS analysis showed that patients with cSLE with any of these gene variants had a similar average PRS as control individuals. RD SNVs were observed in a small proportion of cSLE and carriers of these RD SNVs had a PRS similar to healthy individuals, suggesting the importance of rare coding heterozygous variants in driving disease risk in a subset of children with SLE.

Epstein-Barr virus (EBV) infection constitutes a prerequisite for multiple sclerosis (MS) development, and cross-reactivity between EBV nuclear antigen 1 (EBNA1) and anoctamin-2 (ANO2) antibodies was previously demonstrated in persons with MS (pwMS). Here, we show that ANO2-specific CD4+ T cells are more frequent in pwMS. Immunization of SJL/J mice with ANO2 or EBNA1 led to cross-reactive CD4+ T cell and antibody responses. ANO2 pre-immunization led to exacerbated experimental autoimmune encephalomyelitis (EAE), an effect mediated by CD4+ T cells, as confirmed by adoptive transfer experiments. T cell clones with cross-reactivity to EBNA1 and ANO2 could be isolated from natalizumab-treated pwMS, and sequencing of EBNA1- and ANO2-specific T cell receptors (TCRs) revealed a significant repertoire overlap. We thus report the first mechanistic evidence that EBNA1 CD4+ T cells can target the MS autoantigen ANO2, thereby establishing a link between EBV infection and neuroinflammation.

Psoriasis is a common chronic inflammatory skin disease determined by genetic and environmental factors, resulting in the activation of IL-23/IL-17-mediated immune response, epidermal hyperproliferation, and keratinocyte activation. Long non-coding RNAs (lncRNAs) are non-protein-coding transcripts > 500 nucleotides with diverse regulatory functions; their role in epidermal dysfunction in psoriasis is poorly understood. To identify epidermal transcripts with potential roles in psoriasis, including lncRNAs, we performed RNA sequencing on keratinocytes from psoriasis and healthy skin. We identified 889 differentially expressed lncRNAs, many of which with yet unknown functions. RP11-295G20.2 was identified as a lncRNA significantly induced in psoriasis keratinocytes, and this was verified by qRT-PCR and by single-molecule in situ hybridisation. Analysis of subcellular fractions of epidermis revealed a cytoplasmic localisation in line with results of single molecule in situ hybridisation. We report that RP11-295G20.2 has a skin-enriched expression, and within skin it is mainly expressed in suprabasal epidermal layers. Moreover, RP11-295G20.2 is induced by the key psoriasis cytokine IL-17A and shows a dynamic regulation during keratinocyte differentiation with upregulation during early differentiation and downregulation in the late stage. Knockdown of RP11-295G20.2 in keratinocytes promotes terminal differentiation. Based on our findings, we named RP11-295G20.2 Cytoplasmic Differentiation-Associated Epidermal RNA, CYDAER. In summary, our study provides a comprehensive characterisation of the non-coding RNA landscape of psoriasis keratinocytes and identifies CYDAER as a skin-enriched lncRNA regulating keratinocyte differentiation. Our data suggest that overexpression of CYDAER may contribute to altered differentiation in psoriatic epidermis.

Cyclin A2 is a key regulator of the cell cycle, implicated both in DNA replication and mitotic entry. Cyclin A2 participates in feedback loops that activate mitotic kinases in G2 phase, but why active Cyclin A2-CDK2 during the S phase does not trigger mitotic kinase activation remains unclear. Here, we describe a change in localisation of Cyclin A2 from being only nuclear to both nuclear and cytoplasmic at the S/G2 border. We find that Cyclin A2-CDK2 can activate the mitotic kinase PLK1 through phosphorylation of Bora, and that only cytoplasmic Cyclin A2 interacts with Bora and PLK1. Expression of predominately cytoplasmic Cyclin A2 or phospho-mimicking PLK1 T210D can partially rescue a G2 arrest caused by Cyclin A2 depletion. Cytoplasmic presence of Cyclin A2 is restricted by p21, in particular after DNA damage. Cyclin A2 chromatin association during DNA replication and additional mechanisms contribute to Cyclin A2 localisation change in the G2 phase. We find no evidence that such mechanisms involve G2 feedback loops and suggest that cytoplasmic appearance of Cyclin A2 at the S/G2 transition functions as a trigger for mitotic kinase activation.

How neurons can maintain cellular identity over an entire life span remains largely unknown. Here, we show that maintenance of identity in differentiated dopaminergic and serotonergic neurons is critically reliant on the Polycomb repressive complex 2 (PRC2). Deletion of the obligate PRC2 component, Eed, in these neurons resulted in global loss of H3K27me3, followed by a gradual activation of genes harboring both H3K27me3 and H3K9me3 modifications. Notably, H3K9me3 was lost at these PRC2 targets before gene activation. Neuronal survival was not compromised; instead, there was a reduction in subtype-specific gene expression and a progressive impairment of dopaminergic and serotonergic neuronal function, leading to behavioral deficits characteristic of Parkinson's disease and anxiety. Single-cell analysis revealed subtype-specific vulnerability to loss of PRC2 repression in dopamine neurons of the substantia nigra. Our study reveals that a PRC2-dependent nonpermissive chromatin state is essential to maintain the subtype identity and function of dopaminergic and serotonergic neurons.