Limitations in next-generation sequencing-based genotyping of breast cancer polygenic risk score loci.

Considering polygenic risk scores (PRSs) in individual risk prediction is increasingly implemented in genetic testing for hereditary breast cancer (BC) based on next-generation sequencing (NGS). To calculate individual BC risks, the Breast and Ovarian Analysis of Disease Incidence and Carrier Estimation Algorithm (BOADICEA) with the inclusion of the BCAC 313 or the BRIDGES 306 BC PRS is commonly used. The PRS calculation depends on accurately reproducing the variant allele frequencies (AFs) and, consequently, the distribution of PRS values anticipated by the algorithm. Here, the 324 loci of the BCAC 313 and the BRIDGES 306 BC PRS were examined in population-specific database gnomAD and in real-world data sets of five centers of the German Consortium for Hereditary Breast and Ovarian Cancer (GC-HBOC), to determine whether these expected AFs can be reproduced by NGS-based genotyping. Four PRS loci were non-existent in gnomAD v3.1.2 non-Finnish Europeans, further 24 loci showed noticeably deviating AFs. In real-world data, between 11 and 23 loci were reported with noticeably deviating AFs, and were shown to have effects on final risk prediction. Deviations depended on the sequencing approach, variant caller and calling mode (forced versus unforced) employed. Therefore, this study demonstrates the necessity to apply quality assurance not only in terms of sequencing coverage but also observed AFs in a sufficiently large cohort, when implementing PRSs in a routine diagnostic setting. Furthermore, future PRS design should be guided by the technical reproducibility of expected AFs across commonly used genotyping methods, especially NGS, in addition to the observed effect sizes.

Genetic Architecture of Azoospermia-Time to Advance the Standard of Care.

BACKGROUND: Crypto- and azoospermia (very few/no sperm in the semen) are main contributors to male factor infertility. Genetic causes for spermatogenic failure (SPGF) include Klinefelter syndrome and Y-chromosomal azoospermia factor microdeletions, and CFTR mutations for obstructive azoospermia (OA). However, the majority of cases remain unexplained because monogenic causes are not analysed. OBJECTIVE: To elucidate the monogenic contribution to azoospermia by prospective exome sequencing and strict application of recent clinical guidelines. DESIGN, SETTING, AND PARTICIPANTS: Since January 2017, we studied crypto- and azoospermic men without chromosomal aberrations and Y-chromosomal microdeletions attending the Centre of Reproductive Medicine and Andrology, Münster. OUTCOME MEASUREMENTS AND STATISTICAL ANALYSIS: We performed exome sequencing in 647 men, analysed 60 genes having at least previous limited clinical validity, and strictly assessed variants according to clinical guidelines. RESULTS AND LIMITATIONS: Overall, 55 patients (8.5%) with diagnostic genetic variants were identified. Of these patients, 20 (3.1%) carried mutations in CFTR or ADGRG2, and were diagnosed with OA. In 35 patients (5.4%) with SPGF, mutations in 20 different genes were identified. According to ClinGen criteria, 19 of the SPGF genes now reach at least moderate clinical validity. As limitations, only one transcript per gene was considered, and the list of genes is increasing rapidly so cannot be exhaustive. CONCLUSIONS: The number of diagnostic genes in crypto-/azoospermia was almost doubled to 21 using exome-based analyses and clinical guidelines. Application of this procedure in routine diagnostics will significantly improve the diagnostic yield and clinical workup as the results indicate the success rate of testicular sperm extraction. PATIENT SUMMARY: When no sperm are found in the semen, a man cannot conceive naturally. The causes are often unknown, but genetics play a major role. We searched for genetic variants in a large group of patients and found causal mutations for one in 12 men; these predict the chances for fatherhood.

Bioassay-Guided Fractionation Leads to the Detection of Cholic Acid Generated by the Rare Thalassomonas sp.

Bacterial symbionts of marine invertebrates are rich sources of novel, pharmaceutically relevant natural products that could become leads in combatting multidrug-resistant pathogens and treating disease. In this study, the bioactive potential of the marine invertebrate symbiont Thalassomonas actiniarum was investigated. Bioactivity screening of the strain revealed Gram-positive specific antibacterial activity as well as cytotoxic activity against a human melanoma cell line (A2058). The dereplication of the active fraction using HPLC-MS led to the isolation and structural elucidation of cholic acid and 3-oxo cholic acid. T. actiniarum is one of three type species belonging to the genus Thalassomonas. The ability to generate cholic acid was assessed for all three species using thin-layer chromatography and was confirmed by LC-MS. The re-sequencing of all three Thalassomonas type species using long-read Oxford Nanopore Technology (ONT) and Illumina data produced complete genomes, enabling the bioinformatic assessment of the ability of the strains to produce cholic acid. Although a complete biosynthetic pathway for cholic acid synthesis in this genus could not be determined based on sequence-based homology searches, the identification of putative penicillin or homoserine lactone acylases in all three species suggests a mechanism for the hydrolysis of conjugated bile acids present in the growth medium, resulting in the generation of cholic acid and 3-oxo cholic acid. With little known currently about the bioactivities of this genus, this study serves as the foundation for future investigations into their bioactive potential as well as the potential ecological role of bile acid transformation, sterol modification and quorum quenching by Thalassomonas sp. in the marine environment.

The linear plasmid pSA3239 is essential for the replication of the Streptomyces lavendulae subsp. lavendulae CCM 3239 chromosome.

We previously reported the complete genome of Streptomyces lavendulae subsp. lavendulae CCM 3239, containing the linear chromosome and the large linear plasmid pSA3239. Although the chromosome exhibited replication features characteristic for the archetypal end-patching replication, it lacked the tap/tpg gene pair for two proteins essential for this process. However, this archetypal tpgSa-tapSa operon is present in pSA3239. Complete genomic sequence of the S. lavendulae Del-LP strain lacking this plasmid revealed the circularization of its chromosome with a large deletion of both arms. These results suggest an essential role of pSA3239-encoded TapSa/TpgSa in the end-patching replication of the chromosome.

Streptomonospora litoralis sp. nov., a halophilic thiopeptides producer isolated from sand collected at Cuxhaven beach.

Strain M2T was isolated from the beach of Cuxhaven, Wadden Sea, Germany, in course of a program to attain new producers of bioactive natural products. Strain M2T produces litoralimycin and sulfomycin-type thiopeptides. Bioinformatic analysis revealed a potential biosynthetic gene cluster encoding for the M2T thiopeptides. The strain is Gram-stain-positive, rod shaped, non-motile, spore forming, showing a yellow colony color and forms extensively branched substrate mycelium and aerial hyphae. Inferred from the 16S rRNA gene phylogeny strain M2T affiliates with the genus Streptomonospora. It shows 96.6% 16S rRNA gene sequence similarity to the type species Streptomonospora salina DSM 44593 T and forms a distinct branch with Streptomonospora sediminis DSM 45723 T with 97.0% 16S rRNA gene sequence similarity. Genome-based phylogenetic analysis revealed that M2T is closely related to Streptomonospora alba YIM 90003 T with a digital DNA-DNA hybridisation (dDDH) value of 26.6%. The predominant menaquinones of M2T are MK-10(H6), MK-10(H8), and MK-11(H6) (> 10%). Major cellular fatty acids are iso-C16:0, anteiso C17:0 and C18:0 10-methyl. The polar lipid profile consisted of diphosphatidylglycerol phosphatidyl glycerol, phosphatidylinositol, phosphatidylcholine, phosphatidylethanolamine, three glycolipids, two unknown phospholipids, and two unknown lipids. The genome size of type strain M2T is 5,878,427 bp with 72.1 mol % G + C content. Based on the results obtained from phylogenetic and chemotaxonomic studies, strain M2T (= DSM 106425 T = NCCB 100650 T) is considered to represent a novel species within the genus Streptomonospora for which the name Streptomonospora litoralis sp. nov. is proposed.

Complete genome sequence of Streptomyces cyanogenus S136, producer of anticancer angucycline landomycin A.

Streptomyces cyanogenus S136 is the only known producer of landomycin A (LaA), one of the founding members of angucycline family of aromatic polyketides. LaA displays potent anticancer activities which has made this natural product a target of numerous chemical and cell biological studies. Little is known about the potential of S136 strain to produce other secondary metabolites. Here we report complete genome sequence of LaA producer and how we used this sequence to evaluate for this species its phylogenetic position and diversity of gene clusters for natural product biosynthesis. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s13205-021-02834-4.

The genetic landscape of choroid plexus tumors in children and adults.

BACKGROUND: Choroid plexus tumors (CPTs) are intraventricular brain tumors predominantly arising in children but also affecting adults. In most cases, driver mutations have not been identified, although there are reports of frequent chromosome-wide copy-number alterations and TP53 mutations, especially in choroid plexus carcinomas (CPCs). METHODS: DNA methylation profiling and RNA-sequencing was performed in a series of 47 CPTs. Samples comprised 35 choroid plexus papillomas (CPPs), 6 atypical choroid plexus papillomas (aCPPs) and 6 CPCs plus three recurrences thereof. Targeted TP53 and TERT promotor sequencing was performed in all samples. Whole exome sequencing (WES) and linked-read whole genome sequencing (WGS) was performed in 25 and 4 samples, respectively. RESULTS: Tumors comprised the molecular subgroups "pediatric A" (N=11), "pediatric B" (N=12) and "adult" (N=27). Copy-number alterations mainly represented whole-chromosomal alterations with subgroup-specific enrichments (gains of Chr1, 2 and 21q in "pediatric B" and gains of Chr5 and 9 and loss of Chr21q in "adult"). RNA sequencing yielded a novel CCDC47-PRKCA fusion transcript in one adult choroid plexus papilloma patient with aggressive clinical course; an underlying Chr17 inversion was demonstrated by linked-read WGS. WES and targeted sequencing showed TP53 mutations in 7/47 CPTs (15%), five of which were children. On the contrary, TERT promoter mutations were encountered in 7/28 adult patients (25%) and associated with shorter progression-free survival (log-rank test, p=0.015). CONCLUSION: Pediatric CPTs lack recurrent driver alterations except for TP53, whereas CPTs in adults show TERT promoter mutations or a novel CCDC47-PRKCA gene fusion, being associated with a more unfavorable clinical course.

Class IV Lasso Peptides Synergistically Induce Proliferation of Cancer Cells and Sensitize Them to Doxorubicin.

Heterologous expression of a biosynthesis gene cluster from Amycolatopsis sp. resulted in the discovery of two unique class IV lasso peptides, felipeptins A1 and A2. A mixture of felipeptins stimulated proliferation of cancer cells, while having no such effect on the normal cells. Detailed investigation revealed, that pre-treatment of cancer cells with a mixture of felipeptins resulted in downregulation of the tumor suppressor Rb, making the cancer cells to proliferate faster. Pre-treatment with felipeptins made cancer cells considerably more sensitive to the anticancer agent doxorubicin and re-sensitized doxorubicin-resistant cells to this drug. Structural characterization and binding experiments showed an interaction between felipeptins resulting in complex formation, which explains their synergistic effect. This discovery may open an alternative avenue in cancer treatment, helping to eliminate quiescent cells that often lead to cancer relapse.

High diversity of Vibrio spp. associated with different ecological niches in a marine aquaria system and description of Vibrio aquimaris sp. nov.

The aim of the study was to characterise the diversity and niche-specific colonization of Vibrio spp. in a marine aquaria system by a cultivation-dependent approach. A total of 53 Vibrio spp. isolates were cultured from different ecological niches in a marine aquarium including microplastic (MP) and sandy sediment particles (12 weeks after added sterile to the system), detritus, and the surrounding aquarium water. Based on the 16S rRNA gene sequence phylogeny and multilocus sequence analysis (MLSA) the isolates were assigned to seven different phylotypes. Six phylotypes were identified by high probability to the species level. The highest phylotype diversity was cultured from detritus and water (six out of seven phylotypes), while only two phylotypes were cultured from MP and sediment particles. Genomic fingerprinting indicated an even higher genetic diversity of Vibrio spp. at the strain (genotype) level. Again, the highest diversity of genotypes was recovered from detritus and water while only few partially particle-type specific genotypes were cultured from MP and sediment particles. Phylotype V-2 formed an independent branch in the MLSA tree and could not be assigned to a described Vibrio species. Isolates of this phylotype showed highest 16S rRNA gene sequence similarity to type strains of Vibrio japonicus (98.5%) and Vibrio caribbeanicus (98.4%). A representative isolate, strain THAF100T, was characterised by a polyphasic taxonomic approach and Vibrio aquimaris sp. nov., with strain THAF100T (=DSM 109633T=LMG 31434T=CIP 111709T) as type strain, is proposed as novel species.

Viennamycins: Lipopeptides Produced by a Streptomyces sp.

Extracts from Streptomyces sp. S4.7 isolated from the rhizosphere of edelweiss, an alpine medicinal plant, exhibited activity against Gram-positive bacteria. LC-HRMS analyses of the extracts resulted in the detection of two unknown, structurally related lipopeptides that were assumed to be responsible for the antibiotic activity. LC-MS guided isolation and structure elucidation of viennamycins A and B (1 and 2) by HR-MS/MS, 1D and 2D NMR, and Marfey's analyses revealed them to be novel compounds, with viennamycin A containing cysteic acid, a unique feature for lipopeptides. Tests for antibacterial, antifungal, and cytotoxic activities of purified viennamycins, both with and without divalent cations, did not reveal any bioactivity, suggesting that their biological function, which could not be determined in the tests used, is atypical for lipopeptides. The genome of Streptomyces sp. S4.7 was sequenced and analyzed, revealing the viennamycin biosynthetic gene cluster. Detailed bioinformatics-based analysis of the viennamycin gene cluster allowed elucidation of the biosynthetic pathway for these lipopeptides.

Baikalomycins A-C, New Aquayamycin-Type Angucyclines Isolated from Lake Baikal Derived Streptomyces sp. IB201691-2A.

Natural products produced by bacteria found in unusual and poorly studied ecosystems, such as Lake Baikal, represent a promising source of new valuable drug leads. Here we report the isolation of a new Streptomyces sp. strain IB201691-2A from the Lake Baikal endemic mollusk Benedictia baicalensis. In the course of an activity guided screening three new angucyclines, named baikalomycins A-C, were isolated and characterized, highlighting the potential of poorly investigated ecological niches. Besides that, the strain was found to accumulate large quantities of rabelomycin and 5-hydroxy-rabelomycin, known shunt products in angucyclines biosynthesis. Baikalomycins A-C demonstrated varying degrees of anticancer activity. Rabelomycin and 5-hydroxy-rabelomycin further demonstrated antiproliferative activities. The structure elucidation showed that baikalomycin A is a modified aquayamycin with ß-d-amicetose and two additional hydroxyl groups at unusual positions (6a and 12a) of aglycone. Baikalomycins B and C have alternating second sugars attached, α-l-amicetose and α-l-aculose, respectively. The gene cluster for baikalomycins biosynthesis was identified by genome mining, cloned using a transformation-associated recombination technique and successfully expressed in S. albus J1074. It contains a typical set of genes responsible for an angucycline core assembly, all necessary genes for the deoxy sugars biosynthesis, and three genes coding for the glycosyltransferase enzymes. Heterologous expression and deletion experiments allowed to assign the function of glycosyltransferases involved in the decoration of baikalomycins aglycone.

Complete Genome Sequence of the Cryptophycin-Producing Cyanobacterium Nostoc sp. Strain ATCC 53789.

Nostoc sp. strain ATCC 53789 is a producer of cryptophycins, which are promising anticancer agents. Here, we report the completely sequenced 8.7-Mb genome of Nostoc sp. strain ATCC 53789. The sequence provides insights into the metabolic network of this cyanobacterial strain and illuminates its potential for the biosynthesis of secondary metabolites.

Streptomyces spp. From the Marine Sponge Antho dichotoma: Analyses of Secondary Metabolite Biosynthesis Gene Clusters and Some of Their Products.

Actinomycete bacteria from marine environments represent a potential source for new antibiotics and anti-tumor drugs. Ten strains belonging to the genus Streptomyces isolated from the marine sponge Antho dichotoma collected at the bottom of the Trondheim fjord (Norway) were screened for antibiotic activity. Since only few isolates proved to be bioactive in the conditions tested, we decided to gain an insight into their biosynthetic potential using genome sequencing and analysis. Draft genomes were analyzed for the presence of secondary metabolite biosynthesis gene clusters (BGCs) using antiSMASH software. BGCs specifying both known and potentially novel secondary metabolites were identified, suggesting that these isolates might be sources for new bioactive compounds. The results of this analysis also implied horizontal transfer of several gene clusters between the studied isolates, which was especially evident for the lantibiotic- and thiopeptide-encoding BGCs. The latter implies the significance of particular secondary metabolites for the adaptation of Streptomyces to the spatially enclosed marine environments such as marine sponges. Two bioactive isolates, one showing activity against both yeast and Bacillus subtilis, and one only against yeast were analyzed in details, leading to the identification of cycloheximide, linearmycins, and echinomycins that are presumably responsible for the observed bioactivities.

Prauserella flavalba sp. nov., a novel species of the genus Prauserella, isolated from alkaline soil.

A Gram-strain positive, mycelium-forming actinomycete, YIM 121212T, was isolated from an alkaline soil sample collected in Yunnan province, PR China. Classification using a polyphasic approach indicated that YIM 121212T represents a member of the genus Prauserella, and is closely related to Prauserella coralliicola SCSIO 11529T (99.31 %), Prauserella endophytica SP28S-3T (99.17 %), Prauserella soli 12-833T (97.43 %), Prauserella oleivorans RIPIT (97.03 %), Prauserella marina MS498T (96.74 %), Prauserella rugosa DSM 43194T (96.54 %) and Prauserella muralis 05-Be-005T (95.92 %). Average nucleotide identity values (ANI) of YIM 121212T to P. coralliicola DSM 45821T and P. endophytica CGMCC 4.7182T were 93.1 and 92.8 %, respectively, which were lower than the threshold of 95 %. The digital DNA-DNA hybridization (dDDH) values between YIM 121212T and these two species were 50.8 and 49.9 %, respectively and thus were also well below the cut off value (>70 %) for species delineation. The DNA G+C content of YIM 121212T is 70.8 mol%. Major fatty acids are iso-C16 : 0, iso-C16 : 1H, C16 : 1ω7c/iso-C15 : 0 2OH, C17 : 1ω6c, and C17 : 1ω8c. The predominant menaquinone is MK-9(H4). The polar lipid profile consists of diphosphatidylglycerol (DPG), phosphatidylglycerol (PG), phosphatidylethanolamine (PE), phosphatidylmethylethanolamine (PME), phosphatidylinositol (PI), and phosphatidylinositol mannoside (PIM). The draft genomes were further analyzed for the presence of secondary metabolite biosynthesis (SMB) gene clusters. On the basis of the above observations, YIM121212T can be distinguished from closely related species belonging to the genus Prauserella. Thus, YIM121212T represents a novel species of the genus Prauserella, for which the name Prauserella flavalba sp. nov. is proposed. The type strain is YIM121212T (=CCTCC AA 2013011T=DSM 45973T).

Multi-Omics and Targeted Approaches to Determine the Role of Cellular Proteases in Streptomyces Protein Secretion.

Gram-positive Streptomyces bacteria are profuse secretors of polypeptides using complex, yet unknown mechanisms. Many of their secretory proteins are proteases that play important roles in the acquisition of amino acids from the environment. Other proteases regulate cellular proteostasis. To begin dissecting the possible role of proteases in Streptomyces secretion, we applied a multi-omics approach. We probed the role of the 190 proteases of Streptomyces lividans strain TK24 in protein secretion in defined media at different stages of growth. Transcriptomics analysis revealed transcripts for 93% of these proteases and identified that 41 of them showed high abundance. Proteomics analysis identified 57 membrane-embedded or secreted proteases with variations in their abundance. We focused on 17 of these proteases and putative inhibitors and generated strains deleted of their genes. These were characterized in terms of their fitness, transcriptome and secretome changes. In addition, we performed a targeted analysis in deletion strains that also carried a secretion competent mRFP. One strain, carrying a deletion of the gene for the regulatory protease FtsH, showed significant global changes in overall transcription and enhanced secretome and secreted mRFP levels. These data provide a first multi-omics effort to characterize the complex regulatory mechanisms of protein secretion in Streptomyces lividans and lay the foundations for future rational manipulation of this process.

Complete Genome Sequence of Streptomyces lavendulae subsp. lavendulae CCM 3239 (Formerly "Streptomyces aureofaciens CCM 3239"), a Producer of the Angucycline-Type Antibiotic Auricin.

Streptomyces lavendulae subsp. lavendulae CCM 3239 produces the angucycline antibiotic auricin and was thought to be the type strain of Streptomyces aureofaciens We report the complete genome sequence of this strain, which consists of a linear chromosome and the linear plasmid pSA3239, and demonstrate it to be S. lavendulae subsp. lavendulae.

Comprehensive subcellular topologies of polypeptides in Streptomyces.

BACKGROUND: Members of the genus Streptomyces are Gram-positive bacteria that are used as important cell factories to produce secondary metabolites and secrete heterologous proteins. They possess some of the largest bacterial genomes and thus proteomes. Understanding their complex proteomes and metabolic regulation will improve any genetic engineering approach. RESULTS: Here, we performed a comprehensive annotation of the subcellular localization of the proteome of Streptomyces lividans TK24 and developed the Subcellular Topology of Polypeptides in Streptomyces database (SToPSdb) to make this information widely accessible. We first introduced a uniform, improved nomenclature that re-annotated the names of ~ 4000 proteins based on functional and structural information. Then protein localization was assigned de novo using prediction tools and edited by manual curation for 7494 proteins, including information for 183 proteins that resulted from a recent genome re-annotation and are not available in current databases. The S. lividans proteome was also linked with those of other model bacterial strains including Streptomyces coelicolor A3(2) and Escherichia coli K-12, based on protein homology, and can be accessed through an open web interface. Finally, experimental data derived from proteomics experiments have been incorporated and provide validation for protein existence or topology for 579 proteins. Proteomics also reveals proteins released from vesicles that bleb off the membrane. All export systems known in S. lividans are also presented and exported proteins assigned export routes, where known. CONCLUSIONS: SToPSdb provides an updated and comprehensive protein localization annotation resource for S. lividans and other streptomycetes. It forms the basis for future linking to databases containing experimental data of proteomics, genomics and metabolomics studies for this organism.

Gene panel testing of 5589 BRCA1/2-negative index patients with breast cancer in a routine diagnostic setting: results of the German Consortium for Hereditary Breast and Ovarian Cancer.

The prevalence of germ line mutations in non-BRCA1/2 genes associated with hereditary breast cancer (BC) is low, and the role of some of these genes in BC predisposition and pathogenesis is conflicting. In this study, 5589 consecutive BC index patients negative for pathogenic BRCA1/2 mutations and 2189 female controls were screened for germ line mutations in eight cancer predisposition genes (ATM, CDH1, CHEK2, NBN, PALB2, RAD51C, RAD51D, and TP53). All patients met the inclusion criteria of the German Consortium for Hereditary Breast and Ovarian Cancer for germ line testing. The highest mutation prevalence was observed in the CHEK2 gene (2.5%), followed by ATM (1.5%) and PALB2 (1.2%). The mutation prevalence in each of the remaining genes was 0.3% or lower. Using Exome Aggregation Consortium control data, we confirm significant associations of heterozygous germ line mutations with BC for ATM (OR: 3.63, 95%CI: 2.67-4.94), CDH1 (OR: 17.04, 95%CI: 3.54-82), CHEK2 (OR: 2.93, 95%CI: 2.29-3.75), PALB2 (OR: 9.53, 95%CI: 6.25-14.51), and TP53 (OR: 7.30, 95%CI: 1.22-43.68). NBN germ line mutations were not significantly associated with BC risk (OR:1.39, 95%CI: 0.73-2.64). Due to their low mutation prevalence, the RAD51C and RAD51D genes require further investigation. Compared with control datasets, predicted damaging rare missense variants were significantly more prevalent in CHEK2 and TP53 in BC index patients. Compared with the overall sample, only TP53 mutation carriers show a significantly younger age at first BC diagnosis. We demonstrate a significant association of deleterious variants in the CHEK2, PALB2, and TP53 genes with bilateral BC. Both, ATM and CHEK2, were negatively associated with triple-negative breast cancer (TNBC) and estrogen receptor (ER)-negative tumor phenotypes. A particularly high CHEK2 mutation prevalence (5.2%) was observed in patients with human epidermal growth factor receptor 2 (HER2)-positive tumors.

On the Enigma of Glutathione-Dependent Styrene Degradation in Gordonia rubripertincta CWB2.

Among bacteria, only a single styrene-specific degradation pathway has been reported so far. It comprises the activity of styrene monooxygenase, styrene oxide isomerase, and phenylacetaldehyde dehydrogenase, yielding phenylacetic acid as the central metabolite. The alternative route comprises ring-hydroxylating enzymes and yields vinyl catechol as central metabolite, which undergoes meta-cleavage. This was reported to be unspecific and also allows the degradation of benzene derivatives. However, some bacteria had been described to degrade styrene but do not employ one of those routes or only parts of them. Here, we describe a novel "hybrid" degradation pathway for styrene located on a plasmid of foreign origin. As putatively also unspecific, it allows metabolizing chemically analogous compounds (e.g., halogenated and/or alkylated styrene derivatives). Gordonia rubripertincta CWB2 was isolated with styrene as the sole source of carbon and energy. It employs an assembled route of the styrene side-chain degradation and isoprene degradation pathways that also funnels into phenylacetic acid as the central metabolite. Metabolites, enzyme activity, genome, transcriptome, and proteome data reinforce this observation and allow us to understand this biotechnologically relevant pathway, which can be used for the production of ibuprofen.IMPORTANCE The degradation of xenobiotics by bacteria is not only important for bioremediation but also because the involved enzymes are potential catalysts in biotechnological applications. This study reveals a novel degradation pathway for the hazardous organic compound styrene in Gordonia rubripertincta CWB2. This study provides an impressive illustration of horizontal gene transfer, which enables novel metabolic capabilities. This study presents glutathione-dependent styrene metabolization in an (actino-)bacterium. Further, the genomic background of the ability of strain CWB2 to produce ibuprofen is demonstrated.