Similar to our findings, C-terminal frame shifts and missense mutations of PTEN/MMAC1 have previously been identified in glioblastomas (Ali et al, 1999; Bonneau and Longy, 2000).
sds22 -D2N D199N and sds22 -E163I-L329P and sds22 -77 (C-terminal frame shift) confer cold sensitivity, with accumulation of large budded cells at 14°. sds22 -F117S and sds22 -77 also cause temperature-sensitive growth.
The consequences of such putative alternative splicing events on coding potential include insertion or deletion of between 9 and 28 codons, and limited-range N- or C-terminal frame shifts.
It indicates that N-terminal frame-shift CEBPA mutations, in fact, decisively decrease the amount of wild-type CEBPA protein, whereas the 30-kDa peptide is detectable at a higher amount.
Apart from the original translocation mutation, and a C-terminal frame-shift four nucleotide deletion in DISC1, both of which have the potential to encode protein truncations, numerous nonsynonymous common variants and ultrarare missense mutations in DISC1 have been associated with major mental illness.
The construct containing the GAL4:VP16 fusion to Dnmt2 was created by subcloning the GAL4:VP16 domain from pNT63.10 [21] into a pbGlo2xmyc vector [38] containing the Dnmt2 cDNA to create an N-terminal in frame fusion protein of GAL4:VP16 to Dnmt2, followed by sub-cloning into pWR-Ubq.
Both the cDNAs were cloned into pENTR/D-TOPO vector (Invitrogen), and then transferred into pBA-DC-YFP [ 66] which contains the CaMV 35S promoter and C-terminal in frame YFP, to generate MsTPS1-YFP and MsTPS2-YFP, respectively.
The mSlo3 C-terminal in-frame attached eYFP was made from cloning of two PCR products, 93 bp SalI-NheI fragment from mSlo3 C-terminus and 730 bp NheI-KpnI fragment of eYFP, into SalI-KpnI digested pBF-mSlo3.
The expression of the Drosophila proteins in human HEK 293 cells was monitored by a C-terminal in-frame GFP tag (CG11920 not shown).
