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Identification of genes downstream of Pax6 in the mouse lens using cDNA microarrays
Bharesh K Chauhan1, Nathan A Reed, Weijia Zhang
1Department of Ophthalmology, Albert Einstein College of Medicine, Bronx, New York 10461, USA.
Abstract:
Pax6 is a transcription factor that regulates the development of the visual, olfactory, and central nervous systems, pituitary, and pancreas. Pax6 is required for induction, growth, and maintenance of the lens; however, few direct Pax6 target genes are known. This study was designed to identify batteries of differentially expressed genes in three related systems: 8-week old Pax6 heterozygous lenses, 8-week old Pax6 heterozygous eyes, and transgenic lenses overexpressing PAX6(5a), using high throughput cDNA microarrays containing about 9700 genes. Initially, we obtained almost 400 differentially expressed genes in lenses from mice heterozygous for a Pax6 deletion, suggesting that Pax6 haploinsufficiency causes global changes in the lens transcriptome. Comparisons between the three sets of analyses revealed that paralemmin, molybdopterin synthase sulfurylase, Tel6 oncogene (ETV6), a cleavage-specific factor (Cpsf1) and tangerin A were abnormally expressed in all three experimental models. Semiquantitative reverse transcription (RT)-PCR analysis confirmed that all five of these genes were differentially expressed in Pax-6 heterozygous and Pax6(5a) transgenic lenses. Western blotting and immunohistochemistry demonstrated that paralemmin is found at high levels in the adult lens and confirmed its down-regulation in the Pax6(5a)-transgenic lenses. Collectively, our data provide insights into the genetic programs regulated by Pax6 in the lens.
Insights
Pax6 is crucial for eye development. This study identified five key genes, including paralemmin, whose altered expression in Pax6-deficient or overexpressing lenses reveals new insights into Pax6
Area of Science:
- Developmental Biology
- Genetics
- Ophthalmology
Background:
- Pax6 is a critical transcription factor for nervous system and organ development.
- Pax6 plays a vital role in lens induction, growth, and maintenance.
- Direct Pax6 target genes involved in lens development are not well understood.
Purpose of the Study:
- To identify differentially expressed genes in the lens related to Pax6 function.
- To investigate the impact of Pax6 haploinsufficiency and overexpression on the lens transcriptome.
- To uncover novel Pax6 target genes in lens development.
Main Methods:
- High-throughput cDNA microarrays (approx. 9700 genes) were used.
- Analysis included Pax6 heterozygous lenses and eyes, and transgenic lenses overexpressing PAX6(5a).
- Semiquantitative RT-PCR, Western blotting, and immunohistochemistry were employed for validation.
Main Results:
- Nearly 400 differentially expressed genes were found in Pax6 heterozygous lenses, indicating global transcriptome changes.
- Five genes—paralemmin, molybdopterin synthase sulfurylase, ETV6, Cpsf1, and tangerin A—showed abnormal expression across all models.
- Paralemmin was downregulated in Pax6(5a)-transgenic lenses and highly expressed in adult lenses.
Conclusions:
- Pax6 haploinsufficiency leads to widespread alterations in the lens transcriptome.
- Paralemmin, ETV6, Cpsf1, molybdopterin synthase sulfurylase, and tangerin A are identified as potential direct Pax6 targets in the lens.
- These findings provide new insights into the genetic programs governed by Pax6 during lens development.