scholarly journals Proteomic and Genomic Characterization of a Yeast Model for Ogden Syndrome

2016 ◽  
Author(s):  
Max J. Döerfel ◽  
Han Fang ◽  
Jonathan Crain ◽  
Michael Klingener ◽  
Jake Weiser ◽  
...  
Keyword(s):  
Molecular Chaperones ◽  
Rna Seq ◽  
Stress Tests ◽  
Severe Phenotype ◽  
Cellular Processes ◽  
Hsp70 Family ◽  
Auxiliary Subunit ◽  
Genomic Characterization ◽  
Yeast Model

AbstractNaa10 is a Nα-terminal acetyltransferase that, in a complex with its auxiliary subunit Naa15, co-translationally acetylates the α-amino group of newly synthetized proteins as they emerge from the ribosome. Roughly 40-50% of the human proteome is acetylated by Naa10, rendering this an enzyme with one of the most broad substrate ranges known. Recently, we reported an X-linked disorder of infancy, Ogden syndrome, in two families harboring a c.109T>C (p.Ser37Pro) variant in NAA10. In the present study we performed in-depth characterization of a yeast model of Ogden syndrome. Stress tests and proteomic analyses suggest that the S37P mutation disrupts Naa10 function thereby reducing cellular fitness, possibly due to an impaired functionality of molecular chaperones, Hsp104, Hsp40 and the Hsp70 family. Microarray and RNA-seq revealed a pseudo-diploid gene expression profile in ΔNaa10 cells, likely responsible for a mating defect. In conclusion, the data presented here further support the disruptive nature of the S37P/Ogden mutation and identify affected cellular processes potentially contributing to the severe phenotype seen in Ogden syndrome.

scholarly journals Characterization of Poldip2 knockout mice: avoiding incorrect gene targeting

2021 ◽  
Author(s):  
Bernard Lassègue ◽  
Sandeep Kumar ◽  
Rohan Mandavilli ◽  
Keke Wang ◽  
Michelle Tsai ◽  
...  
Keyword(s):  
Knockout Mice ◽  
Cell Clone ◽  
Gene Trap ◽  
Rna Seq ◽  
Multifunctional Protein ◽  
Cellular Processes ◽  
Depth Study ◽  
Perinatal Lethality ◽  
Knockout Line

AbstractPOLDIP2 is a multifunctional protein whose roles are only partially understood. Our laboratory previously reported physiological studies performed using a mouse gene trap model, which suffered from two limitations: perinatal lethality in homozygotes and constitutive Poldip2 inactivation. To overcome these limitations, we developed a new conditional floxed Poldip2 model. The first part of the present study shows that our initial floxed mice were affected by an unexpected mutation, which was not readily detected by Southern blotting and traditional PCR. It consisted of a 305 kb duplication around Poldip2 with retention of the wild type allele and could be traced back to the original targeted ES cell clone. We offer simple suggestions to rapidly detect similar accidents, which may affect genome editing using both traditional and CRISPR-based methods. In the second part of the present study, correctly targeted floxed Poldip2 mice were generated and used to produce a new constitutive knockout line by crossing with a Cre deleter. In contrast to the gene trap model, many homozygous knockout mice were viable, in spite of having no POLDIP2 expression. To further characterize the effects of Poldip2 ablation in the vasculature, an RNA-seq experiment was performed in constitutive knockout carotid arteries. Results support the involvement of POLDIP2 in multiple cellular processes and provide new opportunities for future in-depth study of its functions.

scholarly journals Characterization of Poldip2 knockout mice: Avoiding incorrect gene targeting

PLoS ONE ◽  
2021 ◽  
Vol 16 (12) ◽  
pp. e0247261
Author(s):  
Bernard Lassègue ◽  
Sandeep Kumar ◽  
Rohan Mandavilli ◽  
Keke Wang ◽  
Michelle Tsai ◽  
...  
Keyword(s):  
Knockout Mice ◽  
Cell Clone ◽  
Gene Trap ◽  
Rna Seq ◽  
Multifunctional Protein ◽  
Cellular Processes ◽  
Depth Study ◽  
Perinatal Lethality ◽  
Knockout Line

POLDIP2 is a multifunctional protein whose roles are only partially understood. Our laboratory previously reported physiological studies performed using a mouse gene trap model, which suffered from three limitations: perinatal lethality in homozygotes, constitutive Poldip2 inactivation and inadvertent downregulation of the adjacent Tmem199 gene. To overcome these limitations, we developed a new conditional floxed Poldip2 model. The first part of the present study shows that our initial floxed mice were affected by an unexpected mutation, which was not readily detected by Southern blotting and traditional PCR. It consisted of a 305 kb duplication around Poldip2 with retention of the wild type allele and could be traced back to the original targeted ES cell clone. We offer simple suggestions to rapidly detect similar accidents, which may affect genome editing using both traditional and CRISPR-based methods. In the second part of the present study, correctly targeted floxed Poldip2 mice were generated and used to produce a new constitutive knockout line by crossing with a Cre deleter. In contrast to the gene trap model, many homozygous knockout mice were viable, in spite of having no POLDIP2 expression. To further characterize the effects of Poldip2 ablation in the vasculature, RNA-seq and RT-qPCR experiments were performed in constitutive knockout arteries. Results show that POLDIP2 inactivation affects multiple cellular processes and provide new opportunities for future in-depth study of its functions.

scholarly journals Proteomic and genomic characterization of a yeast model for Ogden syndrome

Yeast ◽  
2016 ◽  
Vol 34 (1) ◽  
pp. 19-37 ◽  
Author(s):  
Max J. Dörfel ◽  
Han Fang ◽  
Jonathan Crain ◽  
Michael Klingener ◽  
Jake Weiser ◽  
...  
Keyword(s):  
Genomic Characterization ◽  
Yeast Model

Genomic characterization of γ-terpinene synthase from Thymus caespititius

Planta Medica ◽  
2011 ◽  
Vol 77 (12) ◽  
Author(s):  
AS Lima ◽  
B Lukas ◽  
J Novak ◽  
AC Figueiredo ◽  
LG Pedro ◽  
...  
Keyword(s):  
Genomic Characterization

Characterization of CMOS Structures (0.6 µm process) Submitted to HBM and COM ESD Stress Tests

1997 ◽  
Author(s):  
G. Meneghesso ◽  
E. Zanoni ◽  
P. Colombo ◽  
M. Brambilla ◽  
R. Annunziata ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Electrostatic Discharge ◽  
Stress Test ◽  
Complete Characterization ◽  
Stress Tests ◽  
Test Procedures ◽  
Emission Microscopy ◽  
Fast Rise ◽  
Scanning Electron

Abstract In this work, we present new results concerning electrostatic discharge (ESD) robustness of 0.6 μm CMOS structures. Devices have been tested according to both HBM and socketed CDM (sCDM) ESD test procedures. Test structures have been submitted to a complete characterization consisting in: 1) measurement of the tum-on time of the protection structures submitted to pulses with very fast rise times; 2) ESD stress test with the HBM and sCDM models; 3) failure analysis based on emission microscopy (EMMI) and Scanning Electron Microscopy (SEM).

Sign in / Sign up

Export Citation Format

Share Document