Question

Retinitis pigmentosa is a genetic disease that causes the breakdown and loss of retinal cells. Retinitis pigmentosa often starts with decreased night vision and will progress to blindness as retinal cells die.

Several genes have been linked to Retinitis pigmentosa, one of which is GADD. Mice lacking GADD (these mice were experimentally created so that the GADD region of the genome was deleted) have increased expression of non-retinal genes in retinal cells. In other words, in these mutant mice, genes that are not normally expressed in the retina are expressed.  

Crx is a key retinal transcription factor. Crx binds regulatory elements called CBRs (Crx-binding regions). GADD has two CBRs, one immediately proximal to the transcription start site (TSS), and one a few hundred base pairs downstream of the TSS.

You have received DNA from three patients with Retinitis pigmentosa. From the DNA, you sequence the GADD gene and its proximal/core promoter region.   

None of the patients have mutations in the coding region of GADD. All of the mutations you find are in the CBRs. Your findings are below:

• Patient 1 – Mutation in CBR1
• Patient 2 – Mutation in CBR2
• Patient 3 – Mutation in CBR1 and CBR2

Question 7

If you compared histone acetylation in the GADD CBRs between retinal cells and cheek cells would you expect to see more histone acetylation in retinal cells or cheek cells? Explain your answer.

Question 8

How can mutations in non-coding regions cause a change in phenotype (in this case leading to retinitis pigmentosa)? . Note: you do not have to give all possible reasons, explanations of one or two ways mutations in non-coding regions can have phenotypic effects is sufficient.

Question 9

Based on the phenotype of the mice lacking GADD, and what we’ve learned about gene expression what type of protein might GADD be? Explain your answer.

Answer 1

Q7. Retinal cells would have more histone acetylation than cheek cells. Acetylation of lysine residues of histone protein leads to loss of positive charge on lysine that binds with DNA, thereby DNA cannot bind and wrap around histone tightly. This loosened up DNA is available for binding of transcription factors and causes enhancement of transcription. Retinal cells with more acetylation on CBR of GADD, will cause transcriptional activation of GADD, causing expression of retinal genes.

Q8. Mutation in non-coding regions of a gene like promoter or enhancers can reduce or abolish the binding of specific transcription factors. These factors or activators bind to very specific sequences on such non-coding regions and modify transcription level. If a crucial binding sequence (one or more nucleotides) on CBR is mutated, Crx transcription factor cannot bind to it anymore, causing a reduction of transcription of GADD. This will lead to loss of expression of retinal genes.

Q9. GADD is most probably a transcription factor. Mice lacking GADD has reduced expression of retinal genes and enhanced expression of non-retinal genes. Thus it can be inferred that GADD increases the expression of retinal genes in normal cases, by activating the transcription of retinal genes. It is a transcription factor for retinal genes.