Biology subjects

Adams, D.

Publications and source records attributed to Adams, D..

26 records · Page 2Linked to original sources

bioRxiv2021

eIF6 rebinding dynamically couples ribosome maturation and translation

Protein synthesis is a cyclical process consisting of translation initiation, elongation, termination and ribosome recycling. The release factors SBDS and EFL1 (both mutated in the leukaemia predisposition disorder Shwachman-Diamond syndrome) license entry of nascent 60S ribosomal subunits into active translation by evicting the anti-association factor eIF6 from the 60S intersubunit face. Here, we show that in mammalian cells, eIF6 holds all free cytoplasmic 60S subunits in a translationally inactive state and that SBDS and EFL1 are the minimal components required to recycle these 60S subunits back into additional rounds of translation by evicting eIF6. Increasing the dose of eIF6 in mice in vivo impairs terminal erythropoiesis by sequestering post-termination 60S subunits in the cytoplasm, disrupting subunit joining and attenuating global protein synthesis. Our data reveal that ribosome maturation and recycling are dynamically coupled by a mechanism that is disrupted in an inherited leukaemia predisposition disorder.

cell biology↗

bioRxiv2021

Collateral damage: Identification and characterisation of spontaneous mutations causing deafness from a targeted knockout programme

Mice carrying targeted mutations are important for investigating gene function and the role of genes in disease, but the process of culturing embryonic stem cells during the making of a targeted allele offers opportunities for spontaneous mutations to arise. Identifying spontaneous mutations relies on the detection of phenotypes segregating independently of targeted alleles, and many phenotypes are easy to miss if not specifically looked for. Here we present data from a large, targeted knockout programme in which mice were analysed through a phenotyping pipeline. Twenty-five lines out of 1311 displayed different deafness phenotypes that did not segregate with the targeted allele. We have identified 8 different mutations causing deafness in 16 of these 25 lines and characterised the resulting phenotypes. Our data show that spontaneous mutations with observable effects on phenotype are a common side effect of intensive breeding programmes, including those underlying targeted mutation programmes.

genetics↗

bioRxiv2021

Germline loss-of-function variants in the base-excision repair gene MBD4 cause a Mendelian recessive syndrome of adenomatous colorectal polyposis and acute myeloid leukaemia

Inherited defects in base-excision repair (BER) predispose to adenomatous polyposis and colorectal cancer (CRC), yet our understanding of this important DNA repair pathway remains incomplete. By combining detailed clinical, histological and molecular profiling, we reveal biallelic germline loss-of-function (LOF) variants in the BER gene MBD4 to predispose to adenomatous polyposis and -uniquely amongst CRC predisposition syndromes- to myeloid neoplasms. Neoplasms from MBD4-deficient patients almost exclusively accumulate somatic CpG>TpG mutations, resembling mutational signature SBS1. MBD4-deficient adenomas harbour mutations in known CRC driver genes, although AMER1 mutations were more common and KRAS mutations less frequent. We did not find an increased risk for colorectal tumours in individuals with a monoallelic MBD4 LOF variant. We suggest that this condition should be termed MBD4-associated neoplasia syndrome (MANS) and that MBD4 is included in testing for the genetic diagnosis of polyposis and/or early-onset AML.

genetics↗

bioRxiv2020

Cell state diversity promotes metastasis through heterotypic cluster formation in melanoma

In melanoma, transcriptional profiling has revealed multiple co-existing cell states, including proliferative versus invasive sub-populations that have been posited to represent a "go or grow" tradeoff. Both of these populations are maintained in tumors, but how they physically interact to promote metastasis is unknown. We demonstrate that these subpopulations form spatially structured heterotypic clusters that cooperate in the seeding of metastasis. We unexpectedly found that INV cells were tightly adherent to each other, and formed clusters with a rim of PRO cells. Intravital imaging demonstrated cooperation between these populations, in which the INV cells facilitated the spread of less metastatic PRO cells. We identified the TFAP2 neural crest transcription factor as a master regulator of both clustering and the PRO/INV states. Our data suggest a framework for the co-existence of these two divergent cell populations, in which differing cell states form heterotypic clusters that promote metastasis via cell-cell cooperation.

cancer biology↗

bioRxiv2020

Hotspot KRAS mutations in brain metastases at the first metastatic recurrence of cutaneous melanoma

IMPORTANCEBrain metastases occur in 60% of patients with advanced melanoma and are a major cause of melanoma-related mortality and morbidity. Although our understanding of the molecular alterations associated with melanoma progression is improving, there are currently no validated biomarkers which might help identify those patients at highest risk of developing brain metastases. OBJECTIVETo examine the somatic mutational and copy-number landscape of brain metastases that develop as the isolated first visceral site of recurrence - "early brain-metastasis" compared to extracranial melanoma metastases. DESIGN, SETTING AND PARTICIPANTSWhole-exome sequencing of 50 tumors from patients undergoing surgical resection of one or more brain metastasis occurring as the first site of visceral relapse were identified from prospectively maintained databases in Sydney, Wellington, New York and Cambridge. Whole exome sequencing analyses allowed mutational profiles to be compared to cutaneous melanomas in The Cancer Genome Atlas (SKCM-TCGA; n=358) and the Memorial Sloan Kettering (SKCM-MSK-IMPACT; n=186) datasets. An external dataset comprising a further 18 patients with surgically resected early brain metastasis from two additional academic centers served as an independent validation cohort. MAIN OUTCOMES AND MEASURESTo assess the frequency of driver mutations in early brain metastasis and their influence on survival. RESULTSIn concordance with the landmark melanoma sequencing studies, we identified mutations in BRAF (21/50, 42%), NRAS (14/50, 28%) and NF1 (11/50, 22%) as the most frequently mutated melanoma driver genes. When compared to the mutational landscape of cutaneous melanomas in TCGA (SKCM-TCGA), KRAS was the most significantly enriched driver gene, with 5/50 (10%) of brain metastases harboring non-synonymous mutations, of which 4/5 (80%) were in the hotspot positions of codons 12 and 61. This was significantly higher than the corresponding frequency of KRAS-mutations within the entire SKCM-TCGA (2% (7/358), p=0.009, Fishers Exact Test) as well as the SKCM-MSK-IMPACT cohort (1.6% (3/186), p=0.016). Variants in KRAS were mutually exclusive from BRAFV600, NRAS and HRAS mutations and were associated with a significantly reduced overall survival from resection of brain metastasis (relative to KRAS-wild type brain metastases) in multivariate Cox proportional hazard models (HR 1.80, 95% CI 1.46-24.89, p=0.013). Mutations in KRAS were also clonal and concordant with extracranial disease, which suggests these mutations are present within the primary tumor CONCLUSIONS AND RELEVANCEOur analysis, the largest to date, suggests that early metastases to the brain (presenting as the first site of visceral relapse) are characterized by significant enrichment of hotspot KRAS mutations, potentially implicating constitutive RAS-driven cellular programs in neurotropic metastatic behavior in these cases. Based on these data, we suggest that screening for KRAS mutations might help identify those patients with primary melanoma at higher risk of brain metastases or poor survival, and could help inform future surveillance strategies. Key PointsO_ST_ABSQuestionC_ST_ABSWhat is the frequency of driver mutations in early melanoma brain metastases? FindingsIn this study of 50 patients with melanoma metastasizing first to the brain, KRAS mutations were the most significantly enriched driver gene (n=5, 10% of patients) when compared to landmark cutaneous melanoma studies. The high KRAS mutation frequency was also observed in an external validation cohort of 18 patients with early brain metastases. Mutations in KRAS were mutually exclusive from mutations in the key RAS signaling genes and conferred a worse overall survival from resection of brain metastasis. MeaningHotspot KRAS mutations could help identify those patients with primary melanoma at higher risk of brain metastases that may benefit from more intensive, protracted surveillance as well as earlier use of adjuvant therapy.

genomics↗

bioRxiv2019

Multi-site clonality analyses uncovers pervasive subclonal heterogeneity and branching evolution across melanoma metastases

Metastatic melanoma carries a poor prognosis despite modern systemic therapies. Understanding the evolution of the disease could help inform patient management. Through whole-genome sequencing of 13 melanoma metastases sampled at autopsy from a treatment naive patient and by leveraging the analytical power of multi-sample analyses, we reveal that metastatic cells may depart the primary tumour very early in the disease course and follow a branched pattern of evolution. Truncal UV-induced mutations that often swamp downstream analyses of heterogeneity, were found to be replaced by APOBEC-associated mutations in the branches of the evolutionary tree. Multi-sample analyses from a further 7 patients confirmed that branched evolution was pervasive, representing an important mode of melanoma dissemination. Our analyses illustrate that combining cancer cell fraction estimates across multiple metastases provides higher resolution phylogenetic reconstructions relative to single sample analyses and highlights the limitations of accurately inferring inter-tumoural heterogeneity from a single biopsy.

genomics↗

bioRxiv2018

No unexpected CRISPR-Cas9 off-target activity revealed by trio sequencing of gene-edited mice

Introduction Introduction Results and Discussion Conclusion Author Contributions Competing Financial Interests 1. gRNA choices and... 2. Zygote injection 3. DNA extraction from... 4. Amplicon sequencing and... 5. Whole Genome Sequencing 6. Probability of detecting... 7. Variant calling, Trio... 8. Intersection of all... 9. Comparison of SNVs... 10. Validation with PCR... 11. Re-analysis of De... Supplementary Figure Supplementary tables References CRISPR-Cas technologies have transformed genome-editing of experimental organisms and have ...

bioengineering↗

bioRxiv2017

Repeat associated mechanisms of genome evolution and function revealed by the Mus caroli and Mus pahari genomes

Understanding the mechanisms driving lineage-specific evolution in both primates and rodents has been hindered by the lack of sister clades with a similar phylogenetic structure having high-quality genome assemblies. Here, we have created chromosome-level assemblies of the Mus caroli and Mus pahari genomes. Together with the Mus musculus and Rattus norvegicus genomes, this set of rodent genomes is similar in divergence times to the Hominidae (human-chimpanzee-gorilla-orangutan). By comparing the evolutionary dynamics between the Muridae and Hominidae, we identified punctate events of chromosome reshuffling that shaped the ancestral karyotype of Mus musculus and Mus caroli between 3 to 6 MYA, but that are absent in the Hominidae. In fact, Hominidae show between four-and seven-fold lower rates of nucleotide change and feature turnover in both neutral and functional sequences suggesting an underlying coherence to the Muridae acceleration. Our system of matched, high-quality genome assemblies revealed how specific classes of repeats can play lineage-specific roles in related species. For example, recent LINE activity has remodeled protein-coding loci to a greater extent across the Muridae than the Hominidae, with functional consequences at the species level such as reproductive isolation. Furthermore, we charted a Muridae-specific retrotransposon expansion at unprecedented resolution, revealing how a single nucleotide mutation transformed a specific SINE element into an active CTCF binding site carrier specifically in Mus caroli. This process resulted in thousands of novel, species-specific CTCF binding sites. Our results demonstrate that the comparison of matched phylogenetic sets of genomes will be an increasingly powerful strategy for understanding mammalian biology.

genomics↗