{"database":"biostudies-arrayexpress","file_versions":[],"scores":null,"additional":{"submitter":["Michał Śmiga"],"organism":["Porphyromonas gingivalis"],"full_dataset_link":["https://www.ebi.ac.uk/biostudies/studies/E-MTAB-16135"],"description":["Porphyromonas gingivalis is a keystone pathogen responsible for oral microbiome dysbiosis, the development of periodontal disease, and may be involved in the development of systemic comorbidities. As a heme auxotroph, P. gingivalis relies on host hemoproteins as its primary sources of heme and iron. Although heme acquisition in P. gingivalis is relatively well characterized, considerably less is known about its pathways for iron uptake and the regulation of iron and heme homeostasis. P. gingivalis encodes four enzymes associated with the heme biosynthesis pathway, namely HemD, HemN, HemG, and HemH. HemH is a ferrochelatase responsible for the final step of heme biosynthesis, catalyzing the insertion of ferrous iron (Fe²⁺) into protoporphyrin IX. In addition, P. gingivalis expresses the Iht system, which plays a role in iron/heme utilization. One of its components, IhtB, is a heme-binding protein homologous to CbiK cobaltochelatases. Based on this homology, IhtB is hypothesized to function as a reverse ferrochelatase, facilitating the removal of iron from heme and thereby contributing to iron acquisition. This study aimed to investigate the effect of ihtB and hemH gene deletions on P. gingivalis gene expression under heme- and iron-replete conditions, to elucidate the roles of IhtB and HemH chelatases in heme and iron homeostasis."],"repository":["biostudies-arrayexpress"],"sample_protocol":["Growth Protocol - P. gingivalis strains were cultured at 37℃ in anaerobic conditions (80% N2, 10% CO2, 10% H2) on Schaedler anaerobe blood agar (ABA) plates supplemented with 5% sheep blood and vitamin K3 (Argenta, Poznań, Poland) for 3-5 days, and then in a liquid basal medium (BM) composed of 3% trypticase soy broth (Becton Dickinson, Sparks, MD, USA), 0.5% yeast extract (Biomaxima), 0.05 mg/L vitamin K3 (Fluka, Munich, Germany), and 0.05% L-cysteine (Sigma-Aldrich, St. Louis, MO, USA). To grow bacteria in iron and heme-rich conditions, BM was supplemented with 7.7 μM hemin chloride (Pol-Aura, Morąg, Poland) (BM+Hm). Bacteria were grown for 3 passages in Bm+Hm medium. In the final passage, a culture was established in BM+Hm at OD₆₀₀ 0.2 and incubated for ~6 h, reaching an OD₆₀₀ of approximately 0.6-0.7.","Sample Collection - After cultivation, cells were harvested from 2 mL cultures by centrifugation (8,000×g, 10 min, 4°C), and the bacterial pellet was resuspended in 20 µL of 75% ethanol, immediately frozen in liquid nitrogen, and stored at -80℃.","Sequencing - The library was checked with Qubit and real-time PCR for quantification and bioanalyzer for size distribution detection. Quantified libraries were pooled and sequenced on Illumina platforms, according to effective library concentration and data amount required.","Library Construction - Firstly, ribosomal RNA was removed from total RNA, followed by ethanol precipitation. After fragmentation, the first strand cDNA was synthesized using random hexamer primers. During the second strand cDNA synthesis, dUTPs were replaced with dTTPs in the reaction buffer. The directional library was ready after end repair, A-tailing, adapter ligation, size selection, USER enzyme digestion, amplification, and purification.","Nucleic Acid Extraction - Total RNA isolation was performed by Novogene (Cambridge, United Kingdom) according to the company standard protocols."],"figure_sub":["Organization","MINSEQE Score","Assays and Data","Processed Data","MAGE-TAB Files"],"data_protocol":["Data Transformation - Raw reads were mapped to the Porphyromonas gingivalis reference genome using Bowtie2, followed by sorting and indexing. Gene expression levels were quantified based on the number of reads aligned to genomic features. To account for differences in gene length and sequencing depth, read counts were normalized using the FPKM method (Fragments Per Kilobase of transcript per Million mapped reads), enabling accurate comparison of transcript abundances across genes and samples."],"omics_type":["Metabolomics","Unknown","Transcriptomics","Genomics","Proteomics"],"instrument_platform":["Illumina NovaSeq X"],"study_type":["RNA-seq of coding RNA"],"species":["Porphyromonas gingivalis"],"pubmed_authors":["Michał Śmiga"],"additional_accession":[]},"is_claimable":false,"name":"Analysis of gene expression in Porphyromonas gingivalis hemH and ihtB deletion mutant strains grown under heme- and iron-replete conditions (RNA-seq analysis)","description":"Porphyromonas gingivalis is a keystone pathogen responsible for oral microbiome dysbiosis, the development of periodontal disease, and may be involved in the development of systemic comorbidities. As a heme auxotroph, P. gingivalis relies on host hemoproteins as its primary sources of heme and iron. Although heme acquisition in P. gingivalis is relatively well characterized, considerably less is known about its pathways for iron uptake and the regulation of iron and heme homeostasis. P. gingivalis encodes four enzymes associated with the heme biosynthesis pathway, namely HemD, HemN, HemG, and HemH. HemH is a ferrochelatase responsible for the final step of heme biosynthesis, catalyzing the insertion of ferrous iron (Fe²⁺) into protoporphyrin IX. In addition, P. gingivalis expresses the Iht system, which plays a role in iron/heme utilization. One of its components, IhtB, is a heme-binding protein homologous to CbiK cobaltochelatases. Based on this homology, IhtB is hypothesized to function as a reverse ferrochelatase, facilitating the removal of iron from heme and thereby contributing to iron acquisition. This study aimed to investigate the effect of ihtB and hemH gene deletions on P. gingivalis gene expression under heme- and iron-replete conditions, to elucidate the roles of IhtB and HemH chelatases in heme and iron homeostasis.","dates":{"release":"2026-03-03T00:00:00Z","modification":"2026-03-03T02:03:23.936Z","creation":"2025-11-14T14:40:42.924Z"},"accession":"E-MTAB-16135","cross_references":{"ENA":["ERP185021"],"EFO":["EFO_0002944","EFO_0004170","EFO_0003789","EFO_0005518","EFO_0003816","EFO_0003738","EFO_0004184"]}}