(Received for publication, October 3, 1995; and in revised form, December 11, 1995)
From the
In the vertebrate retina, a number of proteins involved in
signal transduction are known to be N-terminal acylated with the
unusual 14 carbon fatty acids 14:1n-9 and 14:2n-6. We
have explored possible pathways for producing these fatty acids in the
frog retina by incubation in vitro with candidate precursor
fatty acids bearing radiolabels, including
[H]14:0,
[
H]18:1n-9,
[
H]18:2n-6, and
[
H]18:3n-3. Rod outer segments were
prepared from the radiolabeled retinas for analysis of protein-linked
fatty acids, and total lipids were extracted from the remaining retinal
pellet. Following saponification of extracted lipids, fatty acid
phenacyl esters were prepared and analyzed by high pressure liquid
chromatography (HPLC) with detection by continuous scintillation
counting. Transducin, whose
-subunit (G
) is known
to bear N-terminal acyl chains, was extracted from the rod outer
segments and subjected to SDS-polyacrylamide gel electrophoresis and
fluorography to detect radiolabeled proteins. G
was
also subjected to methanolysis, and the resulting fatty acyl methyl
esters were analyzed by HPLC. The identities of HPLC peaks coinciding
with unsaturated species of both phenacyl esters and methyl esters were
confirmed by reanalyzing them after catalytic hydrogenation. The
results showed that 14:1n-9 can be derived in the retina from
18:1n-9 and 14:2n-6 from 18:2n-6, most
likely by two rounds of
-oxidation, but that neither is produced
in detectable amounts from 14:0. Retroconversion of unsaturated 18
carbon fatty acids to the corresponding 14 carbon species showed
specificity, in that 18:3n-3 was not converted to 14 carbon
fatty acids in detectable amounts. Myristic acid (14:0),
14:1n-9, and 14:2n-6 were all incorporated into
G
. A much less efficient incorporation of
18:1n-9 into G
was also observed, but no
radiolabeling of G
was observed in retinas incubated
with 18:3n-3. Thus, retroconversion by limited
-oxidation
of longer chain unsaturated fatty acids appears to be the most likely
metabolic source of the unusual fatty acids found on the N termini of
signal transducing proteins in the retina.
A number of proteins of both eukaryotic and viral origin are
modified by fatty acylation through an amide linkage to N-terminal
glycine residues (reviewed by James and Olson(1990), McIlhinney (1990),
Towler et al.(1988), Schlesinger(1993), and Gordon et
al.(1991)). This modification, which in the great majority of
cases studied involves the saturated 14 carbon fatty acid myristate
(14:0), has been shown to play an important role in the function of
these proteins (Jones et al., 1990; Linder et al.,
1991; Yonemoto et al., 1993; Wedegaertner et al.,
1995). In vertebrate retinas, the types of N-terminal fatty acids for
proteins involved in signal transduction are strikingly different from
those found in other tissues. These proteins, which include the
-subunit of the G protein transducin (G
) (
)(Johnson et al., 1994; Kokame et al.,
1992; Neubert et al., 1992; Yang and Wensel, 1992), guanylyl
cyclase-activating protein (GCAP) (Palczewski et al., 1994),
recoverin (Dizhoor et al., 1992; Johnson et al.,
1994), and the catalytic subunit of cAMP-dependent protein kinase
(Johnson et al., 1994), are heterogeneously acylated with
frequent occurrence of 14:1n-9 and 14:2n-6 in
addition to 14:0 and 12:0. This unusual pattern appears to reflect
unusual pathways for synthesizing and utilizing fatty acids for
N-terminal acylation in the retina rather than specific characteristics
of the acylated proteins. The ubiquitous protein kinase is exclusively
modified with 14:0 in the brain and heart (Carr et al., 1982;
Johnson et al., 1994) but in the retina shows the same
heterogeneous pattern of fatty acylation as G
. Indeed,
although 12:0 and 14:0 are found in animal tissues, 14:1n-9
and 14:2n-6 are quite rare and have been found in abundance
only in marine mammals (14:1n-9) (Markley, 1960) and the Asian
plant Evodia rutaecarpa (14:2n-6) (Kurono et
al., 1972).
It is conceivable that the two unsaturated fatty
acids could arise from the desaturation of 14:0. However, previously
described -desaturase enzymes prefer long chain
polyunsaturated fatty acids as substrates (Numa et al., 1984),
and
-desaturases have only been found in a few cell
types, such as testicular Sertoli (Oulhaj et al., 1992) and
brain glioma cells (Cook et al., 1991). Alternatively, these
unusual 14-carbon fatty acids might arise by retroconversion from
longer chain fatty acids through partial
-oxidation. For example,
18:1n-9 (oleic acid) or 18:2n-6 (linoleic acid) might
be converted by this route to 14:1n-9 and 14:2n-6,
respectively. Indeed, it has been hypothesized that 14:2n-6 is
generated within the photoreceptor cell in this manner (Hansen, 1993;
Wang and Anderson, 1993). Such partial
-oxidation is
characteristic of peroxisomal metabolism in contrast to mitochondrial
-oxidation, which favors complete degradation to acetyl-CoA
(Schulz, 1991). Retroconversion pathways of this kind have been
demonstrated to convert 13-hydroxy-9,11-octadecadienoic acid (13-OH,
18:2n-6) to 9-hydroxy-5,7-tetradecadienoic acid (13-OH,
14:2n-6) in lymphocytes (Hadjuagapiiou et al., 1990)
and to 18:2n-6 to 14:2n-6 in rat liver peroxisomes
(Baykousheva et al. 1994). In the retina, a similar
retroconversion pathway was found to convert 22:5n-3 to
20:5n-3 (Wang and Anderson, 1993). We describe here
experiments designed to determine if retroconversion pathways can
produce 14:1n-9 and 14:2n-6 in the frog retina and if
they are used for N-terminal fatty acylation of G
.
To test for resistance of
radiolabel of proteins to hydrolysis by hydroxylamine
(NHOH) as evidence of amide linkage, SDS-PAGE was repeated
and gels were treated with NH
OH using a modification of the
methods of Buss et al.(1987) and Olson et al.(1985).
The gels were fixed (30 min) in isopropanol/H
O/HOAc
(25:65:10, v/v/v), washed (45 min) with H
O, soaked (16 h)
in 1 M NH
OH
HCl (pH 6.7), and washed (6 h)
with isopropanol/H
O/HOAc (10:80:10, v/v/v). Gels were
stained, destained, treated with EN
HANCE(TM), dried, and
autoradiographed. To determine the efficiency of thioester-linked fatty
acid removal by NH
OH, we performed the procedure on
rhodopsin, a protein known to carry two thioester linked 16:0 groups
(Papac et al., 1992). Using previously described procedures,
frog retinas were incubated in vitro with 150 µCi of
[9,10(N)-
H]16:0 (American Radiolabeled
Chemicals Inc., St. Louis, MO), and ROS were prepared, stripped of
transducin, and subjected to SDS-PAGE using low reducing conditions (1
h of incubation in sample application buffer containing 2 mM dithiothreitol and 15 µM 2-mercaptoethanol). Gel
slices (n = 4) containing rhodopsin (
50 µg,
estimated by BCA* protein assay kit, Pierce) were saponified in 1 M NaOH at 37 °C for 2 h and acidified with 12 N HCl,
and radioactivity was determined by counting gel slice and hydrolysate
together (BCS mixture/hydrolysate, 15:1, v/v). Identical gels were
subjected to treatment with 1 M NH
OH as described
above; rhodopsin was excised, saponified, and counted. NH
OH
treatment removed 84 ± 2% of the radiolabel from this protein.
Figure 1:
Incubation with
[9,10-H]14:0. A, HPLC elution profile of
FAPE radioactivity for total lipids extracted from frog retinas
incubated with 1 mCi of [
H]14:0. The y axis ([
H] counts/6-s interval) is shown at a
10
reduction of the original scale; the maximum for
[
H]14:0 peak was 41,900. B, Coomassie
Blue-stained electrophoretic gel (a) and corresponding
fluorogram (b) for transducin (G
)
isolated from frog retinas labeled with 1 mCi of
[
H]14:0. Fluorography was performed for 28 days.
G
and G
are the
- and
-subunits of transducin, respectively. The G
(8
kDa) was run off the gel. C, HPLC elution profiles of FAME
radioactivity for G
and G
(control)
isolated from frog retinas labeled with 1 mCi of
[
H]14:0. FAMEs were released from the
SDS-PAGE-purified G
and G
by acidic
methanolysis.
The Coomassie staining pattern (Fig. 1B, a) and fluorogram (Fig. 1B, b) showed intense radiolabeling at a
migration position in SDS-PAGE aligning precisely with G (39 kDa), and there was no detectable radiolabeling of
G
(36 kDa) or any other proteins. HPLC of methanolysis
products (Fig. 1C) confirmed that the radiolabel on
G
was 14:0 (481 ± 74 counts/6 s/10 µg),
along with some minor peaks of uncertain identity. A comparatively
insignificant amount of labeled 14:0 was associated with G
(18 ± 11 counts/6 s/10 µg).
Figure 2:
Incubation with
[9,10-H]18:1n-9. A, HPLC
elution profile of FAPE radioactivity for total lipids from frog
retinas incubated with 1 mCi of
[
H]18:1n-9. The y axis
([
H] counts/6-s interval) is shown at a 10
reduction of the original scale; the maximum for the
[
H]18:1n-9 peak was at 21,322. Also
shown are the HPLC elution profiles of FAPE radioactivity for the
14:1n-9 and 16:1n-9 after being collected separately
and subjected to catalytic hydrogenation. B, Coomassie
Blue-stained electrophoretic gel (a) and corresponding
fluorogram (b) for transducin (G
)
isolated from frog retinas labeled with 1 mCi of
[
H]18:1n-9. Fluorography was performed
for 41 days. G
and G
are the
-
and
-subunits of transducin, respectively. G
(8
kDa) was run off the gel. C, HPLC elution profiles of FAME
radioactivity for G
and G
(control)
isolated from frog retinas labeled with 1 mCi of
[
H]18:1n-9. FAMEs were released from the
SDS-PAGE-purified G
and G
by acidic
methanolysis. Also shown is the HPLC elution profile of FAME
radioactivity for the 14:1n-9 and 18:1n-9 from the
methanolysis of G
, after being collected
simultaneously and subjected to catalytic
hydrogenation.
The Coomassie staining pattern (Fig. 2B, a) and fluorogram (Fig. 2B, b) revealed faint radiolabeling
concentrated at a migration position in SDS-PAGE aligning precisely
with G (39 kDa), with some diffuse radiolabeling
extending into the region for G
(36 kDa). Another area
of diffuse radiolabeling was observed to be focused at a migration
position aligning with a faint doublet protein band above 84 kDa. HPLC
of methanolysis products (Fig. 2C) showed that the
radiolabel on G
was a mixture of 14:1n-9 (108
± 16 counts/6 s/10 µg) and 18:1n-9 (51 ± 6
counts/6 s/10 µg), whereas the radiolabel seen in the region for
G
was only 18:1n-9 (35 ± 9 counts/6
s/10 µg). HPLC of the G
methanolysis products
after simultaneous collection and catalytic hydrogenation (Fig. 2C) confirmed their identities by showing only
two peaks with appropriate retention times for 14:0 and 18:0 within the
predicted proportions (14:0 (63%) and 18:0 (37%)).
Figure 3:
Incubation with
[9,10,12,13-H]18:2n-6. A, HPLC
elution profile of FAPE radioactivity for total lipids from frog
retinas incubated with 120 µCi of
[
H]18:2n-6. The y axis
([
H] counts/6-s interval) of the radioactivity
profile is shown at a 10
reduction of the original scale; the
maximum for the [
H]18:2n-6 peak was at
32,946. Also shown are the HPLC elution profiles of FAPE radioactivity
for the 14:2n-6 and 16:2n-6 after being collected
separately and subjected to catalytic hydrogenation. B,
Coomassie Blue-stained electrophoretic gel (a) and
corresponding fluorogram (b) for transducin
(G
) isolated from frog retinas labeled with
120 µCi of [
H]18:2n-6. Fluorography
was performed for 42 days. G
and G
are the
- and
-subunits of transducin, respectively.
G
(8 kDa) was run off the gel. C, HPLC
elution profiles of FAME radioactivity for G
and
G
(control) isolated from frog retina labeled with 120
µCi of [
H]18:2n-6. FAMEs were
released from SDS-PAGE-purified G
and G
by acidic methanolysis. Also shown is the HPLC elution profile of
FAME radioactivity for the 14:2n-6 from the methanolysis of
G
, after being subjected to catalytic
hydrogenation.
The Coomassie staining pattern (Fig. 3B, a) and fluorogram (Fig. 3B, b)
revealed moderate radiolabeling at a migration position in SDS-PAGE
aligning precisely with G (39 kDa), with faint
radiolabeling also seen aligning with G
(36 kDa). No
other areas of radiolabeling were observed in the fluorogram. HPLC of
methanolysis products (Fig. 3C) revealed that the
radiolabel on G
was only 14:2n-6 (101
± 8 counts/6 s/10 µg), and no detectable radiolabel was
associated with G
. Catalytic hydrogenation and HPLC of
the methanolysis product for G
confirmed its identity
by showing an appropriate shift in retention time to that of 14:0.
Figure 4:
Incubation with
[9,10,12,13,15,16-H]18:3n-3. A,
HPLC elution profile of FAPE radioactivity for total lipids from frog
retinas incubated with 120 µCi of
[
H]18:3n-3. The y axis
([
H] counts/6-s interval) of the radioactivity
profile is shown at a 4
reduction of the original scale; the
maximum for the [
H]18:3n-3 peak was at
7086. Also shown are the HPLC elution profiles of FAPE radioactivity
for the 18:4n-3 after being subjected to catalytic
hydrogenation. B, Coomassie Blue-stained electrophoretic gel (a) and corresponding fluorogram (b) for transducin
(G
) isolated from frog retinas labeled with
120 µCi of [
H]18:3n-3. Fluorography
was performed for 35 days. G
and G
are the
- and
-subunits of transducin, respectively.
G
(8 kDa) was run off the gel. C, HPLC
elution profiles of FAME radioactivity for G
and
G
(control) isolated from frog retina labeled with 120
µCi of [
H]18:3n-3. FAMEs were
released from SDS-PAGE-purified G
and G
by acidic methanolysis.
The Coomassie staining pattern (Fig. 4B, a) and fluorogram (Fig. 4B, b) showed no radiolabeling
corresponding with G (39 kDa), G
(36
kDa), or any other protein observed on SDS-PAGE. HPLC of methanolysis
products (Fig. 4C) revealed that no detectable
radiolabel was associated with either G
or
G
, consistent with the fluorogram results.
Figure 5:
Hydroxylamine treatment of radiolabeled
transducin. Frog retinas were incubated with either 1 mCi of
[9,10-H] 14:0, 1 mCi of
[9,10-
H]18:1n-9, or 120 µCi of
[9,10,12,13-
H]18:2n-6. Radiolabeled
retinas were used to prepare rod outer segments. Transducin was
isolated and run on the same electrophoretic gel. The gel was treated
with 1 M hydroxylamine and subsequently subjected to
fluorography for 44 days. Shown here are the Coomassie Blue-stained
electrophoretic gel sections after hydroxylamine treatment and the
corresponding fluorograms. G
and G
are the
- and
-subunits of transducin, respectively.
G
(8 kDa) was run off the
gel.
Figure 6:
Distribution of desaturation, enlongation,
or retroconversion products in lipid classes derived from retina
membrane pellets. Total lipids obtained from
[H]14:0,
[
H]18:1n-9,
[
H]18:2n-6, and
[
H]18:3n-3 radiolabeled retina membranes
were resolved into phospholipids (PL), free fatty acids (FFA), and triglycerides (TG) using one-dimensional
two-step TLC. Lipid classes were directly counted to determine total
radioactivity. Lipid classes were then saponified, and the resulting
fatty acids were converted to phenacyl esters and chromatographed on
HPLC. The data are represented as percentages calculated by taking the
ratio of radioactivity for the individual lipid classes or fatty acid
species to the total radioactivity in the original total lipid extract
or separated lipid class.
N-terminal fatty acylation of a protein is carried out
cotranslationally by the enzyme myristoylCoA:protein N-myristoyltransferase (Towler et al., 1988 and
Schlesinger, 1993). Human and yeast myristoyltransferases have a high
substrate specificity for 14:0-CoA (Kishore et al., 1991,
1993; Lu et al., 1994). Although 14:2(n-6)-CoA has
not been tested, 14:1(n-9)-CoA is utilized by both
myristoyltransferases at a 3-fold lower catalytic efficiency (V
/K
) than 14:0-CoA
(Kishore et al., 1993). Despite extensive study, no evidence
has been found for myristoyltransferase isozymes with drastically
altered substrate specificity, and supporting this mRNA from the single
copy human gene appears to be identical in all tissues studied (Duronio et al., 1992). Consequently, to successfully compete for
myristoyltransferase, levels of available 14:1n-9 and
14:2n-6 CoAs must be higher than that of 14:0. To attain these
levels, the retina would need to possess special pathways for
generating 14:1n-9 and 14:2n-6, which are apparently
absent or less active in other tissues. However, this does not mean
14:1n-9 and 14:2n-6 would necessarily accumulate in
the retina, because their CoAs might be taken up rapidly by the
myristoyltransferase during active protein synthesis. This hypothesis
is consistent with the lack of detectable (<0.1%) 14:1n-9
and 14:2n-6 in the total lipid pool of the frog and bovine
retina (Chen and Anderson, 1993a; Bartley et al., 1962).
We
first tested whether 14:1n-9 and 14:2n-6 might be
generated by de novo synthesis, because the 14:1n-5
isomer can be synthesized via a -desaturase (Hamosh
and Bitman, 1992; Koletzko et al., 1992), but we found that
14:0 does not serve as a desaturase substrate in the retina and only
16:0 and 18:0 are produced. We next considered that 14:1n-9
and 14:2n-6 might be produced during the
-oxidation
(retroconversion) of long chain unsaturated fatty acids. Our
experiments showed that retroconversion of 18:1n-9 and
18:2n-6 leads to the formation of 14:1n-9 and
14:2n-6, respectively, in the frog retina. Interestingly,
18:1n-9 was only metabolized to 14:1n-9 and
16:1n-9, suggesting a restricted function for this fatty acid
in the retina. Our results showed 18:2n-6 has multiple roles
in the retina; it was retroconverted to 14:2n-6 and
16:2n-6 and also underwent extensive elongation and
desaturation, following the steps toward 20:4n-6 as in liver
(Sprecher, 1972).
The fatty acid composition of frog retinas (Chen
and Anderson, 1993a) suggests that 18:1n-9 and
16:1n-9 are the likely precursors of 14:1n-9, because
they are the only n-9 fatty acids present in reasonable
abundance (12 and 3% of total fatty acid, respectively). In contrast,
multiple n-6 species are present, with 20:4n-6 being
the most abundant (7%) and 18:2n-6 much less so (1%). Because
rat liver peroxisomes are known to convert 20:4n-6 to
14:2n-6 with 18:3n-6 and 16:3n-6
intermediates (Luthria et al., 1995), 20:4n-6 is a
more likely candidate for the major 14:2n-6 precursor in the
retina. Preliminary results from incubation of
[H]20:4n-6 (DuPont NEN) with frog
retinas (n = 2) showed production of
[
H]16:3n-6 (3%) and
[
H]14:2n-6 (2%) with no detectable
[
H]18:3n-6 (data not shown), supporting
this hypothesis.
N-terminal acylated retina proteins do not contain
14:3n-3 (Johnson et al. 1994), even though n-3 polyunsaturated fatty acids are abundant in the vertebrate
retina (Fliesler and Anderson, 1983). We found that metabolism of
[H]18:3n-3 did not produce detectable
[
H]16:3n-3 or
[
H]14:3n-3 retroconversion products and
radiolabel was not incorporated into G
. This result
shows that the retroconversion process in the frog retina has a
selectivity dependent on double bond position, as well as on chain
length. The 18:3n-3 did undergo extensive elongation and
desaturation, following the steps toward 22:6n-3, as described
for liver (Sprecher, 1972), consistent with previous observations in
the frog retina (Wang and Anderson, 1993).
Under the in vitro incubation conditions we used, there was an active uptake and
metabolism for all the radiolabeled fatty acids, as indicated by the
amount of the original starting radioactivity that was incorporated
into the retina glycerolipids ([H]14:0, 12%;
[
H]18:1n-9, 14%;
[
H]18:2n-6, 5%; and
[
H]18:3n-3, 7%). In all incubations, the
phospholipids contained most of the radiolabel, whereas only a small
percentage was incorporated into triglycerides, consistent with
previous studies (Wang and Anderson, 1993; Chen and Anderson, 1993b).
Incorporation into glycerolipids indicates effective conversion to
precursor CoA ester derivatives. Acyl-CoAs are also the substrates for
myristoyltransferase, and labeling of G
with
[
H]14:0 suggested efficient conversion to the CoA
ester.
In addition to incorporation of
[H]14:0,
[
H]14:1n-9, and
[
H]14:2n-6 into G
, we
also noted some labeling with
[
H]18:1n-9. We have not determined
whether this represents N-terminal acylation by myristoyltransferase,
which in yeast can utilize 18:1n-5
2% as effectively as
14:0 (Rudnick et al., 1992), or trace labeling by
thioesterification or some other means. Our ability to incorporate
[
H]14:2n-6 into frog G
is consistent with the data of Johnson et al.(1994),
which shows 100% modification with 14:2n-6. The incorporation
of [
H]14:0 and
[
H]14:1n-9 into G
indicates that at least some G
in frog retina
can be modified with these fatty acids, possibly at levels too low to
be detected by mass spectrometry.
Amide linkage of the
[H]14:0,
[
H]14:1n-9, and
[
H]14:2n-6 to G
was
supported by finding that the radiolabel was resistant to
hydroxylamine, under conditions where 86% removal of rhodopsin's
thioester-linked fatty acid was achieved. Taken together with the mass
spectrometric results of Johnson et al.(1994) and the well
established specificity of myristoyltransferase in yeast and mammals,
this result strongly suggests that the most likely site for attachment
of these fatty acids is the
-amino group of the N-terminal glycine
of frog G
.
Because frog G is
reported to be modified exclusively by 14:2n-6, we
investigated whether radiolabeled 14:2n-6 would directly
incorporate into the protein. We performed in vitro incubations (n = 2) of frog retinas with 20
µCi of [1-
C]14:2n-6 (data not
shown). Analysis of the retina total lipids showed significant chain
elongation of the [
C]14:2n-6 to
[
C]16:2n-6 and
[
C]18:2n-6 (3 and 4% of
[
C]14:2n-6, respectively). Metabolism
of nonradiolabeled 14:2n-6 to 16:2n-6 and
18:2n-6 has been previously noted in rat liver (Sprecher,
1967). Methanolysis of SDS-PAGE-purified G
(19
± 2 µg) and G
(13 ± 1 µg) from
the [
C]14:2n-6 labeled retinas failed
to release any detectable radiolabeled fatty acids. Because the
[
C]14:2n-6 used in our incubations was
of very low specific activity (55-60 Ci/mol), it is possible that
dilution with endogenously produced 14:2n-6 precluded our
detecting radioactivity in protein product. However, it is also
possible that 14:2n-6 supplied directly may not be readily
available as a substrate for myristoyltransferase to incorporate into
G
. It is clearly converted into the necessary chemical
form, CoA ester, as evidenced by chain elongation products and its
incorporation into glycerolipids. However, subcellular
compartmentalization may limit the access of myristoyltransferase and
the nascent G
polypeptide to 14:2n-6 when it
is supplied directly, while allowing free access to the
14:2n-6 pool produced by retroconversion.
The fatty acid
retroconversions seen in our experiments have the limited chain
shortening characteristics associated with peroxisomal -oxidation
(Schulz, 1991), as shown by the lack of production of 12:1n-9
and 12:2n-6 or other short chain fatty acids. In support of
this are electron microscopy studies showing that
Müller and photoreceptor cells of frog retinas
contain significant peroxisome-like organelles (St. Jules et
al., 1992). However, more experiments are necessary to determine
where the observed
-oxidation occurs. Retina peroxisomes may prove
to be more vigorous or less stringent in retroconverting these fatty
acids compared with those from other tissues. Such differences may
account for the absence of 14:1n-9 or 14:2n-6 on
myristoylated liver proteins such as cytochrome b
reductase
(Ozols et al., 1984), even though liver peroxisomes carry out
fatty acid retroconversions.
Although we do not know the functional
significance for N-terminal fatty acylation of photoreceptor proteins
with unsaturated forms of myristate, the retroconversion pathways we
have investigated could play a major role in both normal visual
function and retinal disease states. Congenital defects such as
adrenomyelnoneuropathy (Moser et al., 1987), neonatal
adrenoleukodystrophy (Jaffe et al., 1982), infantile
Refsum's disease (Poll-the et al., 1987), and Zellweger
syndrome (Bowen et al., 1964) are known afflictions where
peroxisomal -oxidation is impaired. The general phenotype of these
diseases is deterioration of nervous system, often involving the
retina. These symptoms may arise in part from impairment of the
biosynthesis of docosahexaenoic acid (22:6n-3) (Martinez et al., 1994), an essential component of neuronal cells
including retina photoreceptors, which requires the peroxisomal based
retroconversion of 24:6n-3 to 22:6n-3 (Voss et
al., 1991). Because N-terminal fatty acylation with
14:1n-9 and 14:2n-6 may be required for proper
function of phototransduction proteins, impairment of 14:1n-9
or 14:2n-6 production could also have a devastating effect on
normal visual function. It is known that many types of retinal
degeneration involve lipidated phototransduction proteins, such as
rhodopsin (autosomal dominant and autosomal recessive retinitis
pigmentosa) (Dryja et al., 1990; Rosenfeld et al.,
1992) and the
-subunit of cGMP phosphodiesterase (PDE
)
(autosomal recessive retinitis pigmentosa) (McLaughlin et al.,
1993). A specific example of defective protein lipidation in retinal
degeneration is choroideremia, whose basis is a mutation in
geranylgeranyl transferase (Seabra et al., 1993), the enzyme
that isoprenylates PDE
and G proteins of the Rab family.
Therefore, genes encoding proteins involved in the pathways required
for heterogeneous fatty acylation of retina protein warrant further
consideration as retinal degeneration candidates.
The work presented in this paper is based on platform presentations given by James C. DeMar, Jr., at the 86th American Oil Chemists Society 1995 Annual Meeting, San Antonio, TX and the Association for Research in Vision and Ophthalmology 1995 Annual Meeting, Ft. Lauderdale, FL.