Evidence for splice transcript variants of TMEM165, a gene involved in CDG

Area of Science:

• Molecular Biology
• Genetics
• Biochemistry

Background:

• Defects in the TMEM165 gene are linked to Type-II Congenital Disorder of Glycosylation, impacting Golgi processes.
• TMEM165 protein, a conserved Ca2+/H+ antiporter, is crucial for Ca2+ and pH homeostasis, primarily in the Golgi apparatus.
• Patient symptoms include psychomotor retardation, osteoporosis, scoliosis, and bone abnormalities.

Purpose of the Study:

• To investigate the existence and functional implications of TMEM165 splice variants.
• To characterize the localization and effects of different TMEM165 protein isoforms on glycosylation.

Main Methods:

• Reverse Transcription Polymerase Chain Reaction (RT-PCR) and quantitative RT-PCR (RT-Q-PCR) were used to analyze TMEM165 splice-transcript variants and mRNA expression in human brain tissues.
• Expression plasmids were utilized to visualize subcellular localization of TMEM165 isoforms.
• Glycosylation function was assessed by analyzing the gel mobility of highly glycosylated proteins in cells overexpressing TMEM165 isoforms.

Main Results:

• The study identified TMEM165 splice-transcript isoforms, specifically Short-Form (SF) and Long-Form (LF) proteins (129 aa and 259 aa, respectively).
• Both SF and LF isoforms localize to the endoplasmic reticulum and exhibit differential effects on glycosylation compared to the wild-type TMEM165 protein (324 aa).
• SF is expressed at low levels in most tissues but is abundant in the brain, forming homodimers, while LF is exclusively found in the temporal lobe.

Conclusions:

• The discovery of multiple TMEM165 splice variants suggests a family of isoforms.
• These TMEM165 isoforms may play a role in the fine-tuning of TMEM165 protein functions and cellular localization.
• Understanding these isoforms is critical for deciphering the molecular mechanisms underlying TMEM165-related disorders.