Document Type

Dissertation - Open Access

Award Date

2019

Degree Name

Doctor of Philosophy (PhD)

Department / School

Animal Science

First Advisor

George Perry

Keywords

Conceptus, Estradiol, Pregnancy, Proteomics, Transcriptomics, Uterus

Abstract

Preovulatory estradiol has been reported to play a crucial role in pregnancy establishment and maintenance, but the mechanism by which estradiol exerts its effects has not been well characterized. Specifically, the interactions between the maternal uterine environment and the developing conceptus can greatly impact pregnancy success or loss. The objective of this dissertation is to determine the effects of preovulatory estradiol exposure on uterine and trophectoderm transcriptomes, and uterine luminal fluid (ULF) protein composition. Beef cows/heifers were synchronized, artificially inseminated (d 0), and grouped into either high (highE2) or low (lowE2) preovulatory estradiol. Uteri were flushed to collect d16 conceptuses either nonsurgically or following slaughter, and endometrial biopsies (n=29) were collected from the ipsilateral uterine horn. Real-Time PCR (RT-PCR) was performed on trophectoderm (TE; n=21) RNA to measure the relative abundance of IFNT, PTGS2, TM4SF1, C3, FGFR2, and GAPDH. Total cellular RNA was extracted from endometrium for RNA sequencing. ULF pools (n=28) for the following groups: highE2/noconceptus, highE2/conceptus, lowE2/noconceptus, and lowE2/conceptus were analyzed using a 2D LC-MS/MS based 8plex iTRAQ method. RT-PCR data were analyzed using the MIXED procedure in SAS. Transcript abundances in the endometrium were quantified using kallisto, differentially expressed genes (DEGs) were determined using DESeq2 (FDR2), and IPA was used for pathway analysis. Scaffold Q+ was used to quantitate peptide and protein identifications in the ULF. There were no differences in mRNA abundances in TE, but there were 432 DEGs among the highE2/conceptus versus lowE2/conceptus groups, 253 were downregulated (CR2, CDH4, TROAP, COL1A2) and 179 were upregulated (PRSS8, FABP3, IDO1, MUC13, CXCL10) in the highE2/conceptus group. There were 48 differentially expressed proteins (DEPs) among the highE2/conceptus and lowE2/conceptus groups (19 upregulated, 29 downregulated in the highE2/conceptus group), 6 of these were differentially expressed (FDR

Library of Congress Subject Headings

Beef cattle -- Embryos.
Beef cattle -- Embryos -- Mortality.
Estradiol.
Estrus.
Pregnancy.
Ovulation.
Uterus.
Proteomics.

Format

application/pdf

Number of Pages

256

Publisher

South Dakota State University

Comments

The following data sets are available as supplemental files:

  1. Protein and peptiy summary (proteinpilot).xlsx
  2. Scaffold analysis- highE2conceptus vs lowE2conceptus.csv
  3. Scaffold analysis-highE2conceptus vs highE2noconceptus.csv
  4. Scaffold analysis-lowE2conceptus vs lowE2noconceptus.csv
  5. Mass spec raw file.sf3
  6. Raw files from LCMS /MS:
    raw data 8plex. zip
    raw data LTQ.zip
  7. RNA-sequencing_transcripts- file-1.xlxs
  8. RNA-sequencing_transcripts- file-2.xlxs
  9. Complete- proteins- abundences-list

protein and peptiy summary (proteinpilot).xlsx (45109 kB)
Protein and peptiy summary (proteinpilot).xlsx

scaffold analysis- highE2conceptus vs lowE2conceptus.csv (216 kB)
Scaffold analysis- highE2conceptus vs lowE2conceptus.csv

scaffold analysis-highE2conceptus vs highE2noconceptus.csv (216 kB)
Scaffold analysis-highE2conceptus vs highE2noconceptus.csv

scaffold analysis-lowE2conceptus vs lowE2noconceptus.csv (216 kB)
Scaffold analysis-lowE2conceptus vs lowE2noconceptus.csv

mass spec raw file.sf3 (145100 kB)
Mass spec raw file.sf3

rawdata_8plex.zip (651670 kB)
Raw files from LCMS / MS: 8plex

rawdata_LTQ.zip (558439 kB)
Raw files from LCMS / MS: LTQ

RNA-sequencing_transcripts- file-1.xlsx (2494 kB)
Complete list of transcripts from RNA sequencing for the conceptus versus noconceptus comparison

RNA-sequencing_transcripts- file-2.xlsx (1818 kB)
Complete list of transcripts from RNA sequencing for the highE2/noconceptus versus highE2/conceptus comparison and lowE2/noconceptus versus lowE2/conceptus comparison

Complete- proteins- abundences-list.xlsx (457 kB)
complete list of proteins and their abundances for the highE2/ conceptus versus lowE2/conceptus, highE2/conceptus versus highE2/noconceptus, and the lowE2/conceptus versus lowE2/noconceptus comparisons

Included in

Beef Science Commons

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Rights Statement

In Copyright