BRAT1 deficiency causes increased glucose metabolism and mitochondrial malfunction

Abstract

BackgroundBRAT1 (BRCA1-associated ATM activator 1) interacts with both BRCA1, ATM and DNA-PKcs, and has been implicated in DNA damage responses. However, based on our previous results, it has been shown that BRAT1 may be involved in cell growth and apoptosis, besides DNA damage responses, implying that there are undiscovered functions for BRAT1.MethodsUsing RNA interference against human BRAT1, we generated stable BRAT1 knockdown cancer cell lines of U2OS, Hela, and MDA-MA-231. We tested cell growth properties and in vitro/in vivo tumorigenic potentials of BRAT1 knockdown cells compared to control cells. To test if loss of BRAT1 induces metabolic abnormalities, we examined the rate of glycolysis, ATP production, and PDH activity in both BRAT1 knockdown and control cells. The role of BRAT1 in growth signaling was determined by the activation of Akt/Erk, and SC79, Akt activator was used for validation.ResultsBy taking advantage of BRAT1 knockdown cancer cell lines, we found that loss of BRAT1 expression significantly decreases cell proliferation and tumorigenecity both in vitro and in vivo. Cell migration was also remarkably lowered when BRAT1 was depleted. Interestingly, glucose uptake and production of mitochondrial ROS (reactive oxygen species) are highly increased in BRAT1 knockdown HeLa cells. Furthermore, both basal and induced activity of Akt and Erk kinases were suppressed in these cells, implicating abnormality in signaling cascades for cellular growth. Consequently, treatment of BRAT1 knockdown cells with Akt activator can improve their proliferation and reduces mitochondrial ROS concentration.ConclusionsThese findings suggest novel roles of BRAT1 in cell proliferation and mitochondrial functions.Electronic supplementary materialThe online version of this article (doi:10.1186/1471-2407-14-548) contains supplementary material, which is available to authorized users.

BRAT1‐related disease—identification of a patient without early lethality

“…Disruption of its nuclear localization has been shown to be one consequence of a truncating variant [Puffenberger et al, 2012]. A more recent report implicates BRAT1 in apoptosis as well as cellular growth and proliferation, possibly through disruptions in mitochondrial function [So and Ouchi, 2014].…”

“…Recent studies have also implicated it as a regulator of cell growth and apoptosis [So and Ouchi, 2011;So and Ouchi, 2014].…”

PACS-2 mediates the ATM and NF-κB-dependent induction of anti-apoptotic Bcl-xL in response to DNA damage

“…This biphasic PACS-2/ATM interaction is similar to that reported for the interaction between ATM and BRAT1/BAAT1, which, similar to PACS-2, has roles in the nucleus and cytoplasm. 39,40 Whether these biphasic interactions impact compartment-or temporalspecific roles of ATM partners in resolving DNA damage is unknown. The marked reduction of Pacs-2 levels by 6-h post IR (Figures 3a-c and 7a) suggests that the role of this multifunctional protein is temporally regulated in thymocytes.…”

Nedd4 Family Interacting Protein 1 (Ndfip1) Is Required for Ubiquitination and Nuclear Trafficking of BRCA1-associated ATM Activator 1 (BRAT1) during the DNA Damage Response

“…There is accumulating evidence that increasing BRAT1 is beneficial for cell survival, most likely through its regulation of ATM phosphorylation (7). Indeed, reduction of BRAT1 leads to reduced cell proliferation and tumorigenesity (16), mirroring the effects of reduced PI3K signaling and cell survival, although the extent of molecular cross-talk between BRAT1 and PI3K remains elusive. Nonetheless, there is gathering evidence that BRAT1 is a vital part of the armamentarium of the cell against apoptosis during the DNA damage response (19).…”

“…Other constructs used were SF-Ndfip1 (Strep-Flag-Ndfip1 in pcDNA3), Ndfip1 in pBiFC-VN 173 , His-Ubiquitin in pcDNA3, Flag-Nedd4-1 in pcDNA3, Flag-Nedd4-2 in pcDNA3, Flag-Itch in pcDNA3, Ub in pBiFC-VN 173 , F-BimL in pBiFC-VN 173 , Flag-BRAT1 and HA-BRAT1 in pcDNA3 (14,15), and BRAT1 shRNA plasmid (16).…”