Snord 3A: a molecular marker and modulator of prion disease progression.

Since preventive treatments for prion disease require early identification of subjects at risk, we searched for surrogate peripheral markers characterizing the asymptomatic phases of such conditions. To this effect, we subjected blood mRNA from E200K PrP CJD patients and corresponding family members to global arrays and found that the expression of Snord3A, a non-coding RNA transcript, was elevated several times in CJD patients as compared to controls, while asymptomatic carriers presented intermediate Snord3A levels. In the brains of TgMHu2ME199K mice, a mouse model mimicking for E200K CJD, Snord 3A levels were elevated in an age and disease severity dependent manner, as was the case for brains of these mice in which disease was exacerbated by copper administration. Snord3A expression was also elevated in scrapie infected mice, but not in PrP(0/0) mice, indicating that while the expression levels of this transcript may reflect diverse prion etiologies, they are not related to the loss of PrP(C)'s function. Elevation of Snord3A was consistent with the activation of ATF6, representing one of the arms of the unfolded protein response system. Indeed, SnoRNAs were associated with reduced resistance to oxidative stress, and with ER stress in general, factors playing a significant role in this and other neurodegenerative conditions. We hypothesize that in addition to its function as a disease marker, Snord3A may play an important role in the mechanism of prion disease manifestation and progression

Expression of Snord3A in scrapie-infected brain’s.

<p><u>Main picture</u>: Total RNA from brains of scrapie infected mice (6 and 24 months old mice, 4 in each group) and age matched wt controls were amplified for Snord3A by real time PCR as described. Relative RNA expression levels were normalized in reference to UBC. Relative Snord3A expression level in scrapie infected mice is indicated by fold change versus age-matched controls. <u>Insert</u>: Immunoblotting of PrP^Sc in brains of infected mice (2 young, two old). (** P value <0.001).</p

Differential gene expression profile of E200KCJD patients and carriers compared with non carriers control.

<p>Differential gene expression profile of E200KCJD patients and carriers compared with non carriers control.</p

Identification of Snord3A and Aldh1A1 as disease specific markers.

<p>a: Gene expression heat map representing the levels of transcripts expressed in blood from E200KCJD patients, carriers and non-carriers control. b: Snord3A expression level in microarrays of patients and healthy mutation carriers as compared to non-carrier controls. C: Aldh1A1 expression level in patients and healthy mutation carriers as compared to non-carrier controls (P value <0.005).</p

Elevation of Snord3aA expression in Tg MHu2M E199K mice brains is disease dependent.

<p>Total RNA extracted from brains of TgMHu2ME199K (tg), PrP ^0/0 (ko) and wt mice, od designated ages were reverse transcribed into cDNA and amplified by Real Time PCR as described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0054433#s4" target="_blank">Materials and methods</a>. Each group comprised 4–7 mice. Gene expression changes are depicted in relevance to those measured at 3 months old wt mice. Relative expression levels were normalized in reference to UBC. (** P value <0.001).</p

Elevation of Snord3A expression in Tg MHu2M E199K MEFs and in copper treated mice.

<p>Total RNA from brains of TgMHu2ME199K and wild-type mice as described, as well as from designated MEFs were amplified for Snord3A by Real time PCR. Relative RNA expression levels were normalized in reference to UBC and <i>β-actin</i> (Respectively). a: Scheme of accumulation of PrP and MEF survival during copper treatment (as described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0054433#pone.0054433-Canello2" target="_blank">[32]</a>). b: Snord3A levels in designated MEFs (wt: cells from wt mice; tg: cells from TgMHu2ME199K mice; KO: cells from PrP^0/0 mice. c: Snord3A levels in TgMHu2ME199K and wild-type mice brain after 75 days of copper treatment versus age-matched control (** P value <0.001).</p

Multiplex families with epilepsy: success of clinical and molecular genetic characterization

Objective