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In situ localization of globin messenger RNA formation. I. During mouse fetal liver development
Abstract:
Globin mRNA levels in 11-15-day mouse fetal liver cells have been estimated by in situ hybridization of a highly labeled DNA copy (cDNA) of adult globin messenger RNAs (mRNAs) (globin cDNA) to fixed preparations of cells. Under the conditions employed, no significant in situ hybridization occurred to lymphoma cells (L 51787), mouse L cells, or hepatocytes; whereas reticulocytes from phenyl hydrazine-treated mice showed extensive in situ hybridization. The proportion of fetal liver cells showing predominantly cytoplasmic in situ hybridization increased from about 30% at the 11th day of development to 80-85% by days 13-15. Unlike more mature cells, proerythroblasts did not show in situ hybridization, except to a slight extent at later stages of development. These studies therefore indicate that globin mRNAs begin to accumulate during or shortly after the proerythroblastbasophilic erythroblast transition. The fact that certain immature erythroid cells from 14-day fetal liver contain substantial amounts of globin mRNAs has been confirmed by comparing the hybridization in solution of globin cDNA to cytoplasmic RNA extracted from total fetal liver cells or from immature erythroid cells obtained by treatment of fetal liver cells with an antiserum raised against erythrocytes.
Insights
Globin messenger RNA (mRNA) accumulates in mouse fetal liver cells during erythroid development, starting around the proerythroblast stage. This accumulation is crucial for red blood cell formation.
Area of Science:
- Molecular Biology
- Developmental Biology
- Hematopoiesis
Background:
- Globin mRNA is essential for hemoglobin synthesis in red blood cells.
- Understanding the timing of globin mRNA accumulation is key to studying erythropoiesis.
Purpose of the Study:
- To determine when globin mRNA levels increase during mouse fetal liver cell development.
- To identify the specific cell stages involved in globin mRNA accumulation.
Main Methods:
- In situ hybridization using a globin complementary DNA (cDNA) probe on fixed fetal liver cells.
- Quantification of cells with cytoplasmic globin mRNA signal at different developmental days.
- Solution hybridization comparing globin cDNA to RNA from total and purified immature erythroid cells.
Main Results:
- Globin mRNA was detected in fetal liver cells, with signal increasing from 30% at day 11 to 80-85% by days 13-15.
- Proerythroblasts showed minimal to no globin mRNA hybridization, indicating accumulation begins after this stage.
- Hybridization studies confirmed substantial globin mRNA in immature erythroid cells.
Conclusions:
- Globin mRNA accumulation initiates during or shortly after the proerythroblast to basophilic erythroblast transition.
- Immature erythroid cells in fetal liver contain significant amounts of globin mRNA.
- This study provides insights into the temporal regulation of globin gene expression during erythropoiesis.