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Haemoglobin is a key molecule for oxygen transport in vertebrates. It exhibits remarkable gene diversity in teleost fishes, reflecting adaptation to various aquatic environments. In this study, we present the dynamic evolution of haemoglobin subunit genes based on a comparison of high-quality genome assemblies of 24 vertebrate species, including 17 teleosts (of which six are cichlids). Our findings indicate that teleost genomes contain a range of haemoglobin genes, from as few as five in fugu to as many as 43 in salmon, with the latter being the largest repertoire found in vertebrates. We find evidence that the teleost ancestor had at least four Hbα and three or four Hbβ subunit genes, and that the current gene diversity emerged during teleost radiation, driven primarily by (tandem) gene duplications, genome compaction, and rearrangement dynamics. We provide insights into the genomic organisation of haemoglobin clusters in different teleost species. We further show that the evolution of paralogous rhbdf1 genes flanking both teleost clusters (LA and MN) supports the hypothesis for the origin of the LA cluster by rearrangement within teleosts, rather than by the teleost specific whole-genome duplication. We specifically focus on cichlid fishes, where adaptation to low oxygen environment plays role in species diversification. Our analysis of six cichlid genomes, including Pungu maclareni from the Barombi Mbo crater lake, for which we sequenced a representative genome, reveals 18-32 copies of the Hb genes, and elevated rates of non-synonymous substitutions compared to other teleosts. Overall, this work facilitates a deeper understanding of how haemoglobin genes contribute to the adaptive potential of teleosts., (© 2024 The Author(s). Molecular Ecology published by John Wiley & Sons Ltd.)

Background: Delineating base-resolution breakpoints of complex rearrangements is crucial for an accurate clinical understanding of pathogenic variants and for carrier screening within family networks or the broader population. However, despite advances in genetic testing using short-read sequencing (SRS), this task remains costly and challenging., Methods: This study addresses the challenges of resolving missing disease-causing breakpoints in complex genomic disorders with suspected homozygous rearrangements by employing multiple long-read sequencing (LRS) strategies, including a novel and efficient strategy named nanopore-based rapid acquisition of neighboring genomic regions (NanoRanger). NanoRanger does not require large amounts of ultrahigh-molecular-weight DNA and stands out for its ease of use and rapid acquisition of large genomic regions of interest with deep coverage., Findings: We describe a cohort of 16 familial cases, each harboring homozygous rearrangements that defied breakpoint determination by SRS and optical genome mapping (OGM). NanoRanger identified the breakpoints with single-base-pair resolution, enabling accurate determination of the carrier status of unaffected family members as well as the founder nature of these genomic lesions and their frequency in the local population. The resolved breakpoints revealed that repetitive DNA, gene regulatory elements, and transcription activity contribute to genome instability in these novel recessive rearrangements., Conclusions: Our data suggest that NanoRanger greatly improves the success rate of resolving base-resolution breakpoints of complex genomic disorders and expands access to LRS for the benefit of patients with Mendelian disorders., Funding: M.L. is supported by KAUST Baseline Award no. BAS/1/1080-01-01 and KAUST Research Translation Fund Award no. REI/1/4742-01., Competing Interests: Declaration of interests A US provisional patent application (application no. 63/588,122) based on methods described in this paper has been filed by KAUST, in which M.L. and Y.Z. are listed as inventors. F.S.A. is on the advisory board of Med., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

The genomes of living lungfishes can inform on the molecular-developmental basis of the Devonian sarcopterygian fish-tetrapod transition. We de novo sequenced the genomes of the African (Protopterus annectens) and South American lungfishes (Lepidosiren paradoxa). The Lepidosiren genome (about 91 Gb, roughly 30 times the human genome) is the largest animal genome sequenced so far and more than twice the size of the Australian (Neoceratodus forsteri) 1 and African 2 lungfishes owing to enlarged intergenic regions and introns with high repeat content (about 90%). All lungfish genomes continue to expand as some transposable elements (TEs) are still active today. In particular, Lepidosiren's genome grew extremely fast during the past 100 million years (Myr), adding the equivalent of one human genome every 10 Myr. This massive genome expansion seems to be related to a reduction of PIWI-interacting RNAs and C2H2 zinc-finger and Krüppel-associated box (KRAB)-domain protein genes that suppress TE expansions. Although TE abundance facilitates chromosomal rearrangements, lungfish chromosomes still conservatively reflect the ur-tetrapod karyotype. Neoceratodus' limb-like fins still resemble those of their extinct relatives and remained phenotypically static for about 100 Myr. We show that the secondary loss of limb-like appendages in the Lepidosiren-Protopterus ancestor was probably due to loss of sonic hedgehog limb-specific enhancers., (© 2024. The Author(s), under exclusive licence to Springer Nature Limited.)

Background: Perivascular epithelioid cell tumours (PEComas) are soft tissue tumours. These neoplasms belong to the family of mesenchymal tumours, which include angiomyolipomas, clear-cell sugar tumours of the lung, and PEComas not otherwise specified (NOS). The probability of a perivascular epithelioid cell tumour (PEComa) occurring in the uterus is low, and the incidence, diagnosis, treatment, and outcomes of such tumours are still unclear., Case Presentation: A 51-year-old woman presented a 4-year history of natural menopause. An intrauterine mass was detected via ultrasound examination; the mass showed a tendency to increase but caused no symptoms. The levels of tumour markers were within the normal range. Pathological analysis of the curettage revealed perivascular epithelioid differentiation of the endometrial tumour. Consequently, a laparoscopic total hysterectomy with bilateral adnexectomy was performed. No distant metastasis was detected via whole-body positron emission computed tomography (PETCT) after the operation. Fluorescence in situ hybridization (FISH) revealed no TFE3 gene rearrangement. Next-generation sequencing of bone and soft tissue revealed negative TSC1/2 and TP53 expression. No recurrence or metastasis was observed during the 18-month follow-up period., Conclusion: PEComa of the gynecologic tract is a rare and challenging entity. Diffuse HMB-45 expression, TSC alterations and TFE3 rearrangement are characteristic of uterine PEComas. Surgical resection is the first choice. Genetic testing is helpful for determining the nature of the mass and for choosing targeted therapy. Further research is needed to establish treatment protocols., (© 2024. The Author(s).)

Background: Deletion or duplication in the DMD gene is one of the most common causes of Duchenne and Becker muscular dystrophy (DMD/BMD). However, the pathogenicity of complex rearrangements involving DMD, especially segmental duplications with unknown breakpoints, is not well understood. This study aimed to evaluate the structure, pattern, and potential impact of rearrangements involving DMD duplication., Methods: Two families with DMD segmental duplications exhibiting phenotypical differences were recruited. Optical genome mapping (OGM) was used to explore the cryptic pattern of the rearrangements. Breakpoints were validated using long-range polymerase chain reaction combined with next-generation sequencing and Sanger sequencing., Results: A multi-copy duplication involving exons 64-79 of DMD was identified in Family A without obvious clinical symptoms. Family B exhibited typical DMD neuromuscular manifestations and presented a duplication involving exons 10-13 of DMD. The rearrangement in Family A involved complex in-cis tandem repeats shown by OGM but retained a complete copy (reading frame) of DMD inferred from breakpoint validation. A reversed insertion with a segmental repeat was identified in Family B by OGM, which was predicted to disrupt the normal structure and reading frame of DMD after confirming the breakpoints., Conclusions: Validating breakpoint and rearrangement pattern is crucial for the functional annotation and pathogenic classification of genomic structural variations. OGM provides valuable insights into etiological analysis of DMD/BMD and enhances our understanding for cryptic effects of complex rearrangements., (© 2024. The Author(s).)

Background: Recent studies have revealed atypical features in the plastomes of the family Cactaceae, the largest lineage of succulent species adapted to arid and semi-arid regions. Most plastomes sequenced to date are from short-globose and cylindrical cacti, while little is known about plastomes of epiphytic cacti. Published cactus plastomes reveal reduction and complete loss of IRs, loss of genes, pseudogenization, and even degeneration of tRNA structures. Aiming to contribute with new insights into the plastid evolution of Cactaceae, particularly within the tribe Rhipsalideae, we de novo assembled and analyzed the plastomes of Lepismium cruciforme and Schlumbergera truncata, two South American epiphytic cacti., Methods and Results: Our data reveal many gene losses in both plastomes and the first loss of functionality of the trnT-GGU gene in Cactaceae. The trnT-GGU is a pseudogene in L. cruciforme plastome and appears to be degenerating in the tribe Rhipsalideae. Although the plastome structure is conserved among the species of the tribe Rhipsalideae, with tribe-specific rearrangements, we mapped around 200 simple sequence repeats and identified nine nucleotide polymorphism hotspots, useful to improve the phylogenetic resolutions of the Rhipsalideae. Furthermore, our analysis indicated high gene divergence and rapid evolution of RNA editing sites in plastid protein-coding genes in Cactaceae., Conclusions: Our findings show that some characteristics of the Rhipsalideae tribe are conserved, such as plastome structure with IRs containing only the ycf2 and two tRNA genes, structural degeneration of the trnT-GGU gene and ndh complex, and lastly, pseudogenization of rpl33 and rpl23 genes, both plastid translation-related genes., (© 2024. The Author(s), under exclusive licence to Springer Nature B.V.)

Several reports of concurrent MYC, BCL2, BCL6, and CCND1 rearrangements in high-grade B-cell lymphoma (HGBL) have been recently described. Herein, we aimed to delineate the scope of this entity through a review of HGBL with a "quadruple-hit" genetic profile identified at our institution. We performed a retrospective review (2015-2023) at our institution of B-cell lymphoma (BCL) cases that were evaluated with concurrent MYC, BCL2, and BCL6 break-apart and IGH::MYC and IGH::CCND1 dual-color dual-fusion fluorescence in situ hybridization studies. Of 203 cases meeting inclusion criteria, 2 (1%) with a quadruple-hit genetic profile were identified. Case 1 represented a 59-year-old female with widespread lymphadenopathy and a diagnosis of HGBL who exhibited primary refractoriness to dose-adjusted etoposide, prednisone, vincristine, cyclophosphamide, doxorubicin, and rituximab (DA-EPOCH-R) chemotherapy. Case 2 represented a 58-year-old male with mediastinal and abdominal lymphadenopathy and a diagnosis of large BCL who died from disease after 1 cycle of DA-EPOCH-R chemotherapy. Similarly, a literature review of 7 previously reported cases of HGBL with a quadruple-hit profile also demonstrated aggressive disease behavior. Our study adds 2 new cases to the rarely encountered quadruple-hit HGBL, and a brief meta-analysis of the 9 available cases indicates aggressive disease behavior conferred by this constellation of genetic events., (© The Author(s) 2024. Published by Oxford University Press on behalf of American Society for Clinical Pathology. All rights reserved. For commercial re-use, please contact reprints@oup.com for reprints and translation rights for reprints. All other permissions can be obtained through our RightsLink service via the Permissions link on the article page on our site—for further information please contact journals.permissions@oup.com.)

Neuroepithelial tumors with fusion of PLAGL1 or amplification of PLAGL1/PLAGL2 have recently been described often with ependymoma-like or embryonal histology respectively. To further evaluate emerging entities with PLAG-family genetic alterations, the histologic, molecular, clinical, and imaging features are described for 8 clinical cases encountered at St. Jude (EWSR1-PLAGL1 fusion n = 6; PLAGL1 amplification n = 1; PLAGL2 amplification n = 1). A histologic feature observed on initial resection in a subset (4/6) of supratentorial neuroepithelial tumors with EWSR1-PLAGL1 rearrangement was the presence of concurrent ependymal and ganglionic differentiation. This ranged from prominent clusters of ganglion cells within ependymoma/subependymoma-like areas, to interspersed ganglion cells of low to moderate frequency among otherwise ependymal-like histology, or focal areas with a ganglion cell component. When present, the combination of ependymal-like and ganglionic features within a supratentorial neuroepithelial tumor may raise consideration for an EWSR1-PLAGL1 fusion, and prompt initiation of appropriate molecular testing such as RNA sequencing and methylation profiling. One of the EWSR1-PLAGL1 fusion cases showed subclonal INI1 loss in a region containing small clusters of rhabdoid/embryonal cells, and developed a prominent ganglion cell component on recurrence. As such, EWSR1-PLAGL1 neuroepithelial tumors are a tumor type in which acquired inactivation of SMARCB1 and development of AT/RT features may occur and lead to clinical progression. In contrast, the PLAGL2 and PLAGL1 amplified cases showed either embryonal histology or contained an embryonal component with a significant degree of desmin staining, which could also serve to raise consideration for a PLAG entity when present. Continued compilation of associated clinical data and histopathologic findings will be critical for understanding emerging entities with PLAG-family genetic alterations., (© 2024. The Author(s).)

Chromosome rearrangements may distort 3D chromatin architectures and thus change gene regulation, yet how 3D chromatin structures evolve in insects is largely unknown. Here, we obtain chromosome-level genomes for four butterfly species, Graphium cloanthus, Graphium sarpedon, Graphium eurypylus with 2n = 30, 40, and 60, respectively, and Papilio bianor with 2n = 60. Together with large-scale Hi-C data, we find that inter-chromosome rearrangements very rarely disrupted the pre-existing 3D chromatin structure of ancestral chromosomes. However, some intra-chromosome rearrangements changed 3D chromatin structures compared to the ancestral configuration. We find that new TADs and subTADs have emerged across the rearrangement sites where their adjacent compartments exhibit uniform types. Two intra-chromosome rearrangements altered Rel and lft regulation, potentially contributing to wing patterning differentiation and host plant choice. Notably, butterflies exhibited chromatin loops between Hox gene cluster ANT-C and BX-C, unlike Drosophila. Our CRISPR-Cas9 experiments in butterflies confirm that knocking out the CTCF binding site of the loops in BX-C affected the phenotypes regulated by Antp in ANT-C, resulting in legless larva. Our results reveal evolutionary patterns of insect 3D chromatin structures and provide evidence that 3D chromatin structure changes can play important roles in the evolution of traits., (© 2024. The Author(s).)

To make a multicellular organism, genes need to be transcribed at the right developmental stages and in the right tissues. DNA sequences termed 'enhancers' are crucial to achieve this. Despite concerted efforts, the exact mechanisms of enhancer activity remain elusive. Mixed lineage leukemia (MLL or KMT2A) rearrangements (MLLr), commonly observed in cases of acute lymphoblastic leukemia (ALL) and acute myeloid leukemia, produce novel in-frame fusion proteins. Recent work has shown that the MLL-AF4 fusion protein drives aberrant enhancer activity at key oncogenes in ALL, dependent on the continued presence of MLL-AF4 complex components. As well as providing some general insights into enhancer function, these observations may also provide an explanation for transcriptional heterogeneity observed in MLLr patients., Competing Interests: Declaration of Competing Interest T.A.M. is a paid consultant for and shareholder in Dark Blue Therapeutics Ltd., (Copyright © 2024 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Background: Despite the Caridean shrimps' vast species richness and ecological diversity, controversies persist in their molecular classification. Within Caridea, the Pandalidae family exemplifies significant taxonomic diversity. As of June 25, 2023, GenBank hosts only nine complete mitochondrial genomes (mitogenomes) for this family. The Plesionika genus within Pandalidae is recognized as polyphyletic. To improve our understanding of the mitogenome evolution and phylogenetic relationships of Caridea, this study introduces three novel mitogenome sequences from the Plesionika genus: P .  ortmanni , P . izumiae and P . lophotes ., Methods: The complete mitochondrial genomes of three Plesionika species were sequenced utilizing Illumina's next-generation sequencing (NGS) technology. After assembling and annotating the mitogenomes, we conducted structural analyses to examine circular maps, sequence structure characteristics, base composition, amino acid content, and synonymous codon usage frequency. Additionally, phylogenetic analysis was performed by integrating existing mitogenome sequences of true shrimp available in GenBank., Results: The complete mitogenomes of the three Plesionika species encompass 37 canonical genes, comprising 13 protein-coding genes (PCGs), 22 transfer RNAs (tRNAs), two ribosomal RNAs (rRNAs), and one control region (CR). The lengths of these mitogenomes are as follows: 15,908 bp for P . ortmanni , 16,074 bp for P . izumiae and 15,933 bp for P . lophotes . Our analyses extended to their genomic features and structural functions, detailing base composition, gene arrangement, and codon usage. Additionally, we performed selection pressure analysis on the PCGs of all Pandalidae species available in Genbank, indicating evolutionary purification selection acted on the PCGs across Pandalidae species. Compared with the ancestral Caridea, translocation of two tRNA genes, i.e., trnP or trnT , were found in the two newly sequenced Plesionika species- P. izumiae and P. lophotes . We constructed a phylogenetic tree of Caridea using the sequences of 13 PCGs in mitogenomes. The results revealed that family Pandalidae exhibited robust monophyly, while genus Plesionika appeared to be a polyphyletic group., Conclusions: Gene rearrangements within the Pandalidae family were observed for the first time. Furthermore, a significant correlation was discovered between phylogenetics of the Caridea clade and arrangement of mitochondrial genes. Our findings offer a detailed exploration of Plesionika mitogenomes, laying a crucial groundwork for subsequent investigations into genetic diversity, phylogenetic evolution, and selective breeding within this genus., Competing Interests: The authors declare there are no competing interests., (©2024 Sun et al.)

Background: Lysine [K] methyltransferase 2A (KMT2A, previously known as MLL) gene rearrangements are common in acute leukemias of various lineages and are associated with features such as chemotherapy resistance and rapid relapse. KMT2A::CBL is a rare fusion of unknown pathogenesis generated by a unique interstitial deletion of chromosome 11 that has been reported across a wide age range in both acute myeloid leukemia (AML) and acute lymphoblastic leukemia (ALL) patients. The leukemogenic effect of the KMT2A::CBL rearrangement and its association with clinical prognosis have not been well clarified., Methods and Results: We report the case of a 64-year-old female who was diagnosed with acute monoblastic leukemia (M5a) and who acquired the rare KMT2A::CBL fusion. The patient received multiple cycles of therapy but did not achieve remission and eventually succumbed to severe infection and disease progression. Additionally, we characterized the predicted KMT2A-CBL protein structure in this case to reveal the underlying leukemogenic mechanisms and summarized reported cases of hematological malignancies with KMT2A::CBL fusion to investigate the correlation of gene rearrangements with clinical outcomes., Conclusions: This report provides novel insights into the leukemogenic potential of the KMT2A::CBL rearrangement and the correlation between gene rearrangements and clinical outcomes., (© 2024. The Author(s).)

Synthetic Chromosome Rearrangement and Modification by LoxP-mediated Evolution (SCRaMbLE) is a promising tool to study genomic rearrangements. However, the potential of SCRaMbLE to study genomic rearrangements is currently hindered, because a strain containing all 16 synthetic chromosomes is not yet available. Here, we construct SparLox83R, a yeast strain containing 83 loxPsym sites distributed across all 16 chromosomes. SCRaMbLE of SparLox83R produces versatile genome-wide genomic rearrangements, including inter-chromosomal events. Moreover, when combined with synthetic chromosomes, SCRaMbLE of hetero-diploids with SparLox83R leads to increased diversity of genomic rearrangements and relatively faster evolution of traits compared to hetero-diploids only with wild-type chromosomes. Analysis of the SCRaMbLEd strain with increased tolerance to nocodazole demonstrates that genomic rearrangements can perturb the transcriptome and 3D genome structure and consequently impact phenotypes. In summary, a genome with sparsely distributed loxPsym sites can serve as a powerful tool for studying the consequence of genomic rearrangements and accelerating strain engineering in Saccharomyces cerevisiae., (© 2024. The Author(s).)

Purpose: To report genetic characteristics and associated risk of chromosomal breaks due to chromosomal rearrangements in large samples., Methods: MicroSeq, a technique that combines chromosome microdissection and next-generation sequencing, was used to identify chromosomal breakpoints. Long-range PCR and Sanger sequencing were used to precisely characterize 100 breakpoints in 50 ABCR carriers., Results: In addition to the recurrent regions of balanced rearrangement breaks in 8q24.13, 11q11.23, and 22q11.21 that had been documented, we have discovered a 10-Mb region of 12q24.13-q24.3 that could potentially be a sparse region of balanced rearrangement breaks. We found that 898 breakpoints caused gene disruption and a total of 188 breakpoints interrupted genes recorded in OMIM. The percentage of breakpoints that disrupted autosomal dominant genes recorded in OMIM was 25.53% (48/188). Fifty-four of the precisely characterized breakpoints had 1-8-bp microhomologous sequences., Conclusion: Our findings provide a reference for the evaluation of the pathogenicity of mutations in related genes that cause protein truncation in clinical practice. According to the characteristics of breakpoints, non-homologous end joining and microhomology-mediated break-induced replication may be the main mechanism for ABCRs formation., (© 2023. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)