Gestational Age Dependence of the Maternal Circulating Long Non-Coding RNA Transcriptome During Normal Pregnancy Highlights Antisense and Pseudogene Transcripts.

Erica L Kleinbrink,Nardhy Gomez-Lopez,Roberto Romero,Leonard Lipovich,Donghong Ju,Adi L Tarca,Anton-Scott Goustin,Bogdan Done

doi:10.3389/fgene.2021.760849

Abstract

In the post-genomic era, our understanding of the molecular regulators of physiologic and pathologic processes in pregnancy is expanding at the whole-genome level. Longitudinal changes in the known protein-coding transcriptome during normal pregnancy, which we recently reported (Gomez-Lopez et al., 2019), have improved our definition of the major operant networks, yet pregnancy-related functions of the non-coding RNA transcriptome remain poorly understood. A key finding of the ENCODE (Encyclopedia of DNA Elements) Consortium, the successor of the Human Genome Project, was that the human genome contains approximately 60,000 genes, the majority of which do not encode proteins. The total transcriptional output of non-protein-coding RNA genes, collectively referred to as the non-coding transcriptome, is comprised mainly of long non-coding RNA (lncRNA) transcripts (Derrien et al., 2012). Although the ncRNA transcriptome eclipses its protein-coding counterpart in abundance, it has until recently lacked a comprehensive, unbiased, genome-scale characterization over the timecourse of normal human pregnancy. Here, we annotated, characterized, and selectively validated the longitudinal changes in the non-coding transcriptome of maternal whole blood during normal pregnancy to term. We identified nine long non-coding RNAs (lncRNAs), including long intergenic non-coding RNAs (lincRNAs) as well as lncRNAs antisense to or otherwise in the immediate vicinity of protein-coding genes, that were differentially expressed with advancing gestation in normal pregnancy: AL355711, BC039551 (expressed mainly in the placenta), JHDM1D-AS1, A2M-AS1, MANEA-AS1, NR_034004, LINC00649, LINC00861, and LINC01094. By cross-referencing our dataset against major public pseudogene catalogs, we also identified six transcribed pseudogenes that were differentially expressed over time during normal pregnancy in maternal blood: UBBP4, FOXO3B, two Makorin (MKRN) pseudogenes (MKRN9P and LOC441455), PSME2P2, and YBX3P1. We also identified three non-coding RNAs belonging to other classes that were modulated during gestation: the microRNA MIR4439, the small nucleolar RNA (snoRNA) SNORD41, and the small Cajal-body specific ncRNA SCARNA2. The expression profiles of most hits were broadly suggestive of functions in pregnancy. These time-dependent changes of the non-coding transcriptome during normal pregnancy, which may confer specific regulatory impacts on their protein-coding gene targets, will facilitate a deeper molecular understanding of pregnancy and lncRNA-mediated molecular pathways at the maternal-fetal interface and of how these pathways impact maternal and fetal health.

Highlights

Transcriptomes flux and change dramatically across both space and time throughout all organismal developmental programs, and during the execution of all normal cellular and organismal functions as well as in disease processes
We previously reported that transcripts that are differentially expressed with gestational age in normal pregnancy were enriched in genes located on chromosome 14
We found that expression profiles of the Timecourse Differentially Expressed (TDE) lncRNAs and lincRNAs modulated during gestation in normal pregnancy are typically tissue-specific, as is expected of functional lncRNAs, and were broadly consistent with pregnancy-associated functions

Summary

Introduction

Transcriptomes flux and change dramatically across both space and time throughout all organismal developmental programs, and during the execution of all normal cellular and organismal functions as well as in disease processes. Microbial genomes in fetal plasma (Witt et al, 2020) were recently detected from umbilical cord blood, highlighting the utility of next-generation sequencing to detect intra-amniotic infection and for personalized diagnostics of highrisk complications of preterm birth. Despite these key advances, little remains known about regulation of the non-coding transcriptome during pregnancy, including small (such as micro) and long non-coding RNAs, and the epigenetic as well as post-transcriptional, cis as well as trans regulatory effects of pregnancy-responsive ncRNAs on the proteins and at the key genomic loci that are functionally relevant to pregnancy and/or its complications. The roles of these genes will subsequently need to be functionally validated with direct follow-up gain- and/or loss-of-function studies of these ncRNAs in primary cell culture or organoid models of direct relevance to pregnancy

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