Identifying unique regulatory elements related to polymorphic imprinting and gestational aging in the placenta

Lead Research Organisation: University of East Anglia
Department Name: Biological Sciences

Abstract

The placenta grows during pregnancy and is attached to the wall of the womb. It links the mother to the baby via the umbilical cord. Its main purpose is to supply the baby with all nutrients and oxygen required for growth, whilst removing waste products and carbon dioxide from the baby's blood. If the placenta does not develop or function correctly, this can lead to pregnancy complications that have huge consequences for the baby, not immediately following delivery, but later in life. Cells are the fundamental unit of complex tissues such as the placenta. Cell phenotype and function are very different between the different cell types. It is the code and activity (referred to as expression) of our genes that are ultimately responsible for defining which cells are produced and how they behave. This is achieved by 'epigenetic' modifications to the DNA. One form of epigenetic regulation is achieved by marking certain regions of the genome with methyl groups that tend to act as a semi-permanent block for gene expression. However, DNA methylation does not act alone, there are different interactive mechanisms that help in epigenetic regulation. In recent years a class of genes, known as imprinted genes, have been shown to be essential for placenta development and function. These uniquely regulated genes are epigenetically turned "on" and expressed solely from the maternal or paternal copy, but not both. The placenta is fascinating as it has a unique epigenetic profile having significantly less DNA methylation compared to all other tissues, which subtly changes during pregnancy. The hierarchical consequences of this unique hypomethylated state on the addition layers of epigenetic information have not been investigated. Furthermore, the placenta harbours hundreds of unique imprinted genes not found in other tissues, the expression of which generally decrease with gestation in a DNA methylation independent manner. This suggests other epigenetic mechanisms are responsible for the observed downregulation. Our understanding of this phenomenon has been hindered by the fact we do not know the mechanisms responsible for healthy aging within the placenta. In this application, we seek to understand how methylation in placenta influences other epigenetic marks at different time point in pregnancy and how this ultimately impacts on imprinted gene expression (Objectives 1.1-1.2). To achieve this, we will utilize, and in some cases improve, the latest technologies to identify which area of the genome are associated with different epigenetic signatures and how this influence development (Objectives 2.1 & 3). Furthermore, since we anticipate variability and cell-type specific signatures within our samples, we will profile isolated single-cells, allowing for us to identify patterns which would not be evident in "bulk" placenta biopsies (Objective 2.2). To ensure maximum chance of success building of our previous experiences, we have established a team that includes researchers responsible for technical development at the UK National Capability in Genomics at the Earlham Institute and computational experts for data analysis. Following the analysis using extremely powerful computers and specialised computer programs (Objective 2.3) we will compare our placenta-derived data across time, and within difference cells- types, to identify regions important for gestation-age related expression changes, especially for imprinted genes, which we will subsequently be remove or alter to determine functionality (Objective 4).

Technical Summary

In this application we propose a series of experiments to better understand the hierarchical placental epigenome across gestation (by assessing two different developmental time points) to determine intra-sample variability and to gain widespread functional insights of potential cis-acting regulatory elements with unique placenta profiles. The data will be generated and assessed in a genome-wide unbiased fashion, allowing us to monitor gestational-dynamics in imprinted gene expression and identify cis-regulatory usage that would account for the widespread down-regulation of expression previously reported.

There is an inter-relationship between DNA methylation and additional layers of epigenetic information essential for genome function. We have previously shown that the placenta is uniquely hypomethylated and that this methylation is dynamic through pregnancy. However, addition epigenetic tiers have largely been uncharacterised in this essential tissue and mechanisms responsible for the global increase in methylation (on the unique hypomethylated background) are unknown. Fascinatingly, transcriptional down-regulation associated with imprinted transcripts at term is independent of methylation, since all imprinted differentially methylated regions maintain faithful allelic profiles, suggesting other epigenetic mechanisms must be responsible.

Main deliverables: (1) We will produced consensus epigenetic maps for early and late placenta samples. Through exploitation of these datasets, we will ultimately identify functional regulatory elements, many of which we anticipate will change during gestation. (2) By performing bioinformatic analyses using trio-SNP information to discriminate alleles, the temporal epigenetic and expression profiles for imprinted genes will be revealed. Since the epigenetic datasets will be produced in an unbiased genome-wide fashion, this will allow novel cis-regulatory elements to be defined and functionally characterised.
 
Description We have now generated the full epigenome maps for two cell types from term placenta samples as well as establish trophoblast and stromal cell lines. We have optimised a MACs protocol for isolating trophoblast and non-trophoblast fractions of the placenta using cell surface markers specific to each cell-type. These were original objectives outlines in the award. We are currently preparing a manuscript describing the protocols which will enable other researchers to reproducibly isolate these cells and to allow future comparison with our epigenetic atlas. In addition, we have identified and characterised several novel placenta-specific gDMRs that originate from the oocyte and looked at the evolutionary conservation of non-canonical imprints in human placenta.
Exploitation Route Having optimised cell isolation protocols, other researchers can enrich specific cell-types from placenta samples. Furthermore, once finished, our epigenome maps will give a greater understanding of this key tissue in health and disease
Sectors Healthcare

Pharmaceuticals and Medical Biotechnology

 
Description Our results have impact within academia, as results from our placenta epigenome project have altered our interpretation of genomic imprinting in humans. This has now been incorporated in the undergraduate and Master's taught course content. Results have also helped develop a novel nuclear-isolation protocol to enrich cell-type specific DNA from frozen samples, something that will be widely beneficial to researchers using archived samples and those collecting samples from remote locations (i.e. this is being utilised by researcher in rural Gambia, Africa). Through wide public dissemination, local Norfolk communities have a deeper understanding of development and pregnancy.
First Year Of Impact 2023
Sector Education
Impact Types Societal

 
Description Design education material for Masters courses at UEA
Geographic Reach Local/Municipal/Regional 
Policy Influence Type Influenced training of practitioners or researchers
Impact Increased participation in MSc dissertation projects in the related research field, as well as enquiries for PhD positions following MSc graduation.
 
Description Member of the International Consensus group for Multi-locus Imprinting Disorders
Geographic Reach Multiple continents/international 
Policy Influence Type Participation in a guidance/advisory committee
Impact The aim of the meeting was to develop a 'road-map' of work leading to a consensus statement on diagnosis and management of MLID. The manuscript is currently being drafted.
 
Description BBSRC NRP DTP PhD fellowship for Keran Chen
Amount £130,000 (GBP)
Funding ID BB/T008717/1 
Organisation Biotechnology and Biological Sciences Research Council (BBSRC) 
Sector Public
Country United Kingdom
Start 08/2022 
End 08/2026
 
Description NIHR Research Capability Funding
Amount £19,000 (GBP)
Organisation Norfolk and Norwich University Hospitals NHS Foundation Trust 
Sector Public
Country United Kingdom
Start 09/2023 
End 01/2024
 
Description NIHR Research Capability Funding
Amount £13,947 (GBP)
Organisation Norfolk and Norwich University Hospitals NHS Foundation Trust 
Sector Public
Country United Kingdom
Start 02/2024 
End 04/2024
 
Description Collaboration to look at imprinting profile in placenta samples with Beckwith-Wiedemann syndorme patients 
Organisation Hospices Civils de Lyon
Country France 
Sector Hospitals 
PI Contribution We have hybridised FFPE-derived DNA samples from archived samples to Illumina EPIC arrays and performed bespoke bioinformatic analysis of both imprinted domains and genome-wide.
Collaborator Contribution The partners have characterised the samples at the histopathological level and supplied samples for downstream molecular characterisation
Impact Assay design for confirmation of aberrant epigenetic profile. Preparation of manuscript for publication
Start Year 2022
 
Description Collaboration to look at the role of MYPT1 variants on NO-induced placenta vessel vasodilation 
Organisation Norfolk and Norwich University Hospital
Country United Kingdom 
Sector Hospitals 
PI Contribution We have processed human placenta tissue samples that have been deposited with the Norwich Research Park Biorepository.
Collaborator Contribution Our partner, Dr Jon Lartey, is consultant gyeancologist at NNUH as is responsible for recruiting, consenting and collecting placenta tissues for downstream processing.
Impact We are generating a collection of extra-embryonic tissues in collaboration with the Norwich Research Park Biorepository. These will be available to all researcher on completion of a samples request form.
Start Year 2022
 
Description DNA methylation profiling in extreme prematures - determining the influence of newborn feeding regimes 
Organisation Norfolk and Norwich University Hospitals NHS Foundation Trust
Country United Kingdom 
Sector Public 
PI Contribution We have performed the bioinformatic analysis of HM450k methylation arrays. Samples were collected at birth, with several follow-up samples to determine lasting epigenetic changes due to milk type and supplementation
Collaborator Contribution Our collaborator, Dr Iglesias-Platas, was responsible for collecting the cohort of samples and obtaining all the associated meta-data
Impact A manuscript is in preparation. Discussions are ongoing for a larger follow-up study
Start Year 2023
 
Description DNA methylation profiling in placenta-derived tumours 
Organisation Imperial College London
Department Faculty of Medicine
Country United Kingdom 
Sector Academic/University 
PI Contribution DNA methylation profiling in placenta-derived tumours: we are performing the bioinformatic analysis of Illumina EPIC array datasets
Collaborator Contribution Our partner and project lead, Geoffrey Maher, has been responsible for collecting these extremely rare samples, performing sample characterisation and generating the dataset.
Impact Once the data is assessed, the work will be submitted for publication
Start Year 2023
 
Description DNA extraction demonstration/experiments with the Camberely Scout Group, Norwich 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach Local
Primary Audience Public/other audiences
Results and Impact The evening consisted of 3 group rotations. We discussed genetics, DNA inheritance and epigenetics to teenagers. This was followed by hands-on DNA extraction from Strawberries.
Year(s) Of Engagement Activity 2024
URL https://www.facebook.com/7thcamberleyscoutgroup/?locale=en_GB
 
Description Invited speaker at European Society of Human Genetics conference 2023 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Postgraduate students
Results and Impact I was an invited speaker at European Society of Human Genetics conference 2023 in Glascow, giving the Opening Session presentation on "Placenta Epigenetics".
Year(s) Of Engagement Activity 2023
URL https://2023.eshg.org/
 
Description Made mini-video for students at Kinsale Infant school, Norwich. 
Form Of Engagement Activity Engagement focused website, blog or social media channel
Part Of Official Scheme? No
Geographic Reach Local
Primary Audience Schools
Results and Impact Made mini-video describing out research (placenta biology: what was out belly button used for?)/virtual guided tour of the lab for year 2 students (7yr-olds) at xxxx school, Norwich. This was part of their Science Week activities.
Year(s) Of Engagement Activity 2024
 
Description Norwich Science Week 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach Regional
Primary Audience Public/other audiences
Results and Impact The Norwich Science festival is a seven day event over half-term. We participated in several events, including (1) talking to children/general public about DNA, (2) talking to general children/public about placenta and pregnancy complications
Year(s) Of Engagement Activity 2024
URL https://norwichsciencefestival.co.uk/
 
Description Organising committee member for the Wellcome Connecting Science Conference on Genomic Imprinting 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Professional Practitioners
Results and Impact This was a hybrid event was the second in the series with over 100 in-person participants and 150 online. This was a three day event that included lectures, discussions and workshop. It primarily aimed to highlight advances in imprinting research and how research can best be translated into clinical benefit. The participants were from multidisciplinary backgrounds (basic research scientists, healthcare professionals and others from different disciplines) and forged many cross-disciplinary collaborations.
Year(s) Of Engagement Activity 2023
URL https://coursesandconferences.wellcomeconnectingscience.org/event/genomic-imprinting-from-biology-to...
 
Description Organising/hosting Pint of Science event 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach Regional
Primary Audience Public/other audiences
Results and Impact Organising/hosting Pint of Science events with local scientists sharing their research around genetics and health
Year(s) Of Engagement Activity 2023,2024
URL https://pintofscience.co.uk/events/norwich
 
Description Royal Norfolk show 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Public/other audiences
Results and Impact We were involved with the UEA stand at the Royal Norfolk show. The stands were aimed at school children informing them about (1) the genetics of taste and smell, (2) STEMM Village exhibition
Year(s) Of Engagement Activity 2023
URL https://www.royalnorfolkshow.co.uk/
 
Description School Visit to (Eaton Junior, Norwich) March 2022 for class discussion on early embryo development. 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach Local
Primary Audience Schools
Results and Impact ~30 pupils participated in an extended classroom visit, which sparked questions and discussion afterwards.
Year(s) Of Engagement Activity 2022
 
Description School Visit to (Eaton Junior, Norwich) March 2023 for class discussion on "Human Development" as part of Science Week 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach Local
Primary Audience Schools
Results and Impact Presented to two separate classes of ~30 pupils an extended classroom demonstration, which sparked questions and discussion afterwards, which was a part of the 2024 Science Week.
Year(s) Of Engagement Activity 2024
 
Description School Visit to (Eaton Junior, Norwich) March 2023 for class discussion on "how the placenta works" as part of Science Week 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach Local
Primary Audience Schools
Results and Impact ~30 pupils participated in an extended classroom demonstration, which sparked questions and discussion afterwards, which was a part of the 2023 Science Week.
Year(s) Of Engagement Activity 2023
 
Description School Visit to (Heathersett Primary, Norwich) June 2023 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach Local
Primary Audience Schools
Results and Impact School Visit to Heathersett for class discussion on "how the placenta works" and "human development" to primary school children aged 6-7yrs.
Year(s) Of Engagement Activity 2023