Question

You have found that genea, which codes for proteina, has been implicated in Autism Spectrum Disorders. As a leader in your field, you would like to learn a few things about this gene and how it influences cellular activity. From what you have read in previously published research, decreased expression of gene results in both increased oxidative stress and inflammation. There are a number of places where those signaling pathways can converge, and you'd like to figure out if Transcriptional Activator 1 (TA1) for inflammatory cytokine TNF-a is that convergence point. Aim 1: Determine if expression of gene is related to activity, or expression, of TA1. How will you test for levels of TA1 and genea? What model will you use? Create a graph that shows that there is an inverse relationship. Now you want to know if you can offset the inflammatory response generated by reduced protein by inhibiting the activity of TA1. Aim 2: Determine if TAl is necessary for inflammatory response following genea knockdown. How will you knockdown genea? How will you measure inflammation? How will you inhibit TA1? Create a graph that shows that inhibiting TA1 reduces inflammation Now the important part: Aim 3: Determine if TA1 inhibition rescues the behavioral phenotype. What behavior will you measure and why? Create a graph that shows that TA1 inhibition rescues the behavioral phenotype in your mouse model.

I am struggling with the first portion of the question (aim #1) more than any other and would like help! Thanks in advance!

Answer 1

The protein levels of ASD patients were examined by western blotting. The gene expression was determined by qRT-PCR. Gene expression analysis identified a 35% decrease in ERβ mRNA expression in the middle frontal gyrus of ASD subjects. In addition, a 38% reduction in aromatase (CYP19A1) mRNA expression was observed in ASD subjects. There will also be significant decreases in ER co-activators that included a 34% decrease in SRC-1, a 77% decrease in CBP, and a 52% decrease in P/CAF mRNA levels in ASD subjects relative to controls.

Oxidative stress has been implicated in the pathogenesis of several inflammatory lung disorders. Oxidants and inflammatory mediators such as tumour necrosis factor-α (TNF-α) activate transcription factors such as nuclear factor-kappaB (NF-κB) and activator protein-1 (AP-1) leading to the expression of pro-inflammatory genes.

To investigate whether the effect of TNF-α on SHBG was via NF-κB in our in vivo system, human SHBG transgenic mice were treated with daily intraperitoneal injections of TNF-α (5 μg) or TNF-α (5 μg) plus the NF-κB inhibitor QNZ (1 μg) for 5 days. The graph is attached.

SHBG levels + A 7 Control TNF a 0 3 5 Days