3 research outputs found
Comparative Nucleotide-Dependent Interactome Analysis Reveals Shared and Differential Properties of KRas4a and KRas4b
The KRAS gene encodes
two isoforms, KRas4a and KRas4b. Differences
in the signaling functions of the two KRas proteins are poorly understood.
Here we report the comparative and nucleotide-dependent interactomes
of KRas4a and KRas4b. Many previously unknown interacting proteins
were identified, with some interacting with both isoforms while others
prefer only one. For example, v-ATPase a2 and eIF2Bδ interact
with only KRas4b. Consistent with the v-ATPase interaction, KRas4b
has a significant lysosomal localization. Comparing WT and constitutively
active G12D mutant KRas, we examined differences in the effector proteins
of the KRas4a and KRas4b. Interestingly, KRas4a binds RAF1 stronger
than KRas4b. Correspondingly, KRas4a can better promote ERK phosphorylation
and anchorage-independent growth than KRas4b. The interactome data
represent a useful resource to understand the differences between
KRas4a and KRas4b and to discover new function or regulation for them.
A similar proteomic approach would be useful for studying numerous
other small GTPases
Comparative Nucleotide-Dependent Interactome Analysis Reveals Shared and Differential Properties of KRas4a and KRas4b
The KRAS gene encodes
two isoforms, KRas4a and KRas4b. Differences
in the signaling functions of the two KRas proteins are poorly understood.
Here we report the comparative and nucleotide-dependent interactomes
of KRas4a and KRas4b. Many previously unknown interacting proteins
were identified, with some interacting with both isoforms while others
prefer only one. For example, v-ATPase a2 and eIF2Bδ interact
with only KRas4b. Consistent with the v-ATPase interaction, KRas4b
has a significant lysosomal localization. Comparing WT and constitutively
active G12D mutant KRas, we examined differences in the effector proteins
of the KRas4a and KRas4b. Interestingly, KRas4a binds RAF1 stronger
than KRas4b. Correspondingly, KRas4a can better promote ERK phosphorylation
and anchorage-independent growth than KRas4b. The interactome data
represent a useful resource to understand the differences between
KRas4a and KRas4b and to discover new function or regulation for them.
A similar proteomic approach would be useful for studying numerous
other small GTPases
HDAC8 Catalyzes the Hydrolysis of Long Chain Fatty Acyl Lysine
The histone deacetylase (HDAC) family
regulates many biological
pathways through the deacetylation of lysine residues on histone and
nonhistone proteins. Mammals have 18 HDACs that are classified into
four classes. Class I, II, and IV are zinc-dependent, while class
III is nicotinamide adenine dinucleotide (NAD<sup>+</sup>)-dependent
lysine deacetylase or sirtuins. HDAC8, a class I HDAC family member,
has been shown to have low deacetylation activity compared to other
HDACs <i>in vitro.</i> Recent studies showed that several
sirtuins, with low deacetylase activities, can actually hydrolyze
other acyl lysine modifications more efficiently. Inspired by this,
we tested the activity of HDAC8 using a variety of different acyl
lysine peptides. Screening a panel of peptides with different acyl
lysine modifications, we found that HDAC8 can catalyze the removal
of acyl groups with 2–16 carbons from lysine 9 of the histone
H3 peptide (H3K9). Interestingly, the catalytic efficiencies (<i>k</i><sub>cat</sub>/<i>K</i><sub>m</sub>) of HDAC8
on octanoyl, dodecanoyl, and myristoyl lysine are several-fold better
than that on acetyl lysine. The increased catalytic efficiencies of
HDAC8 on larger fatty acyl groups are due to the much lower <i>K</i><sub>m</sub> values. T-cell leukemia Jurkat cells treated
with a HDAC8 specific inhibitor, PCI-34051, exhibited an increase
in global fatty acylation compared to control treatment. Thus, the
de-fatty-acylation activity of HDAC8 is likely physiologically relevant.
This is the first report of a zinc-dependent HDAC with de-fatty-acylation
activity, and identification of HDAC8 de-fatty-acylation targets will
help to further understand the function of HDAC8 and protein lysine
fatty acylation