Differential expression pattern of the novel
serine⁄ threonine kinase, STK33, in mice and men
Alejandro O. Mujica
1
*, Bastienne Brauksiepe
1
*, Sigrid Saaler-Reinhardt
2
, Stefan Reuss
3
and
Erwin R. Schmidt
1
1 Institute of Molecular Genetics, Johannes Gutenberg-University, Mainz, Germany
2 Institute of Genetics, Johannes Gutenberg-University, Mainz, Germany
3 Department of Anatomy and Cell Biology, Johannes Gutenberg-University, Mainz, Germany
The serine ⁄ threonine kinase 33 gene (STK33 ⁄ Stk33)
was identified by comparative sequencing of human
chromosome region 11p15.3 and its syntenic region in
mouse chromosome 7 [1,2]. Chromosome 11p15.3 is a
gene-rich region of clinical importance because several
human diseases, including predisposition for some
types of cancer, have been mapped there [3,4]. It has
also been associated with several defects and malig-
nancies, such as the Beckwith–Wiedemann syndrome,
haemoglobinopathies, Long QT syndrome (Ward–
Romano Syndrome), insulin-dependent diabetes mellitus
I, Usher syndrome 1C, T-cell leukaemia, hypoparathy-
roidism and Nieman–Pick disease type A and B
(reviewed in [5]) as well as different types of cancer in
urinary bladder, ovary, testis, breast and lung [6,7].

(Received 6 May 2005; revised 2 August
2005, accepted 3 August 2005)
doi:10.1111/j.1742-4658.2005.04900.x
Serine ⁄ threonine kinase 33 (STK33 ⁄ Stk33) is a recently discovered gene
whose inferred amino acid sequence translation displays characters typical
for a calcium ⁄ calmodulin dependent kinase (CAMK). In this study we ana-
lysed the STK33 ⁄ Stk33 RNA and protein distribution and the localization
of the protein. The STK33 ⁄ Stk33 expression pattern resembles those of
some related members of the CAMK group. STK33 ⁄ Stk33 displays a non-
ubiquitous and, in most tissues, low level of expression. It is highly
expressed in testis, particularly in cells from the spermatogenic epithelia.
Moreover, significant expression is detected in lung epithelia, alveolar
macrophages, horizontal cells in the retina and in embryonic organs such as
heart, brain and spinal cord. A possible role of STK33 ⁄ Stk33 in spermato-
genesis and organ ontogenesis is discussed.
Abbreviations
STK33 ⁄ Stk33, human and mouse serine ⁄ threonine kinase proteins; STK33 ⁄ Stk33: human and mouse serine ⁄ threonine kinase genes; CAMK:
Ca
2+
⁄ calmodulin dependent kinases group; CAMK1 ⁄ Camk1, CAMK2 ⁄ Camk2, CAMK4 ⁄ Camk4: genes for Ca
2+
⁄ calmodulin dependent
kinases I, II and IV from human (in capitals) and mouse; FCS, fetal calf serum; dpp, days postpartum; MTE, multiple tissue expression;
ISH, in situ hybridization; LSM, laser scanning microscopy; Hsa, Homo sapiens; Mmu, Mus musculus.
4884 FEBS Journal 272 (2005) 4884–4898 ª 2005 FEBS
[15–21]. Certain alternative splicing variants of
CAMK2 were found to be expressed preferentially in
tumour cells [22] and the Camk2d isoform is downreg-
ulated in both human and mouse tumour cells [23].
CAMK4 expression has also been associated with epi-

lower total expression than, for example, housekeeping
genes such as GAPDH (16014 human EST ⁄ 1128
mouse EST) or b-actin (15776 ⁄ 4559). STK33 ⁄ Stk33
EST counts are similar to other CAMKs (CAMK1:
133 ⁄ 148, CAMK2A: 141 ⁄ 129, CAMK4:83⁄ 177) and
their distribution suggests a nonubiquitous expression
(Table 1 shows a human ⁄ mouse comparison together
Table 1. Comparison of human and mouse STK33 ⁄ Stk33 expression levels between our data and expression databases.
Organ ⁄ tissue
Hsa STK33 Mmu Stk33 Hsa STK33 Mmu Stk33 Hsa STK33 Mmu Stk33 Hsa STK33 Mmu Stk33
This paper
Positive experiments
UniGene
a
count [32]
Number ⁄ % of clones
EST profile viewer
a
[32]
Transcripts per million
SOURCE Gene
Report
b
[34]
% Normalized
Testis +++ +++ 17 ⁄ 16.0 34 ⁄ 68.0 124 329 10.89 71.57
Lung + + ++ 22 ⁄ 20.7 0 76 0 6.97 –
Fetal lung + – 0 0 – – – –
Heart – – 0 1 ⁄ 2 0 18 – 4.96
Fetal heart + + 0 0 – – – –

ESTs from human lung and testis are most frequently
represented (20.7% and 16.0%, respectively). Embry-
onic and fetal tissues as well as diverse tumour tissues
and cancer cell lines are also represented with several
entries each. Some entries are present from tissues of
the nervous system as well as from auditory and ocular
systems. Prostate and uterus also show single STK33
entries. The EST coverage for Stk33 in the mouse
seems to be more limited. Only testis is very well repre-
sented with 68% of the entries, and interestingly there
are no entries from the lung. In addition to testis, there
are single ESTs from retina, pituitary gland and cir-
cumventricular organs of the brain such as subfornical
organ and area postrema.
In this first survey, we have addressed the expression
of STK33 ⁄ Stk33 focussing on RNA and protein distri-
bution with emphasis on the mouse as an animal
model. Manning and colleagues [8] explained the fail-
ure to detect some novel kinases, despite their similar-
ity to members of the superfamily, with the limited
expression of these proteins. The results presented here
suggest that this may be the case for STK33 and that
its expression pattern also resembles that of members
of the CAMK family of protein kinases. As a step
towards understanding their function, we have
analysed the distribution of STK33 ⁄ Stk33 RNA and
protein as well as the subcellular localization of the
protein.
Results
Presence of STK33 ⁄ Stk33 RNA in mouse and

+
RNA were loaded (lg): heart, 4; intestine,
20; brain, 20; kidney, 6; ovary, 6; testis, 12; lung, 15. Normaliza-
tion was performed by simultaneous hybridization with a probe
from mouse ribosomal protein gene L19. The arrow shows a
possible shorter alternative transcript present in testis. (B) cDNA
dot-blot (MTE, Clontech) hybridization with an STK33 specific
probe, with samples from different regions of nervous system
(NS), heart (He), digestive system (DS), several organs (SO), can-
cer cell lines (Ca), fetal organs (FO) and diverse controls (Co).
The first column with practically no signal corresponds to several
regions of brain (see Fig. S1 for the identity of each dot). (C)
Quantification of the signal obtained in the cDNA dot-blot normal-
ized to the maximal signal in testis. Only results are shown from
tissues with signals higher than the highest value for the negat-
ive controls.
STK33 expression pattern A. O. Mujica et al.
4886 FEBS Journal 272 (2005) 4884–4898 ª 2005 FEBS
The expression pattern of STK33 in human was
investigated by cDNA dot-blot hybridization using the
human multiple tissue expression (MTE, Clontech,
Palo Alto, CA) and a STK33 specific probe. The
cDNA dot-blot contains 76 normalized dots of immo-
bilized cDNA, 61 from adult normal tissues, eight can-
cer cell lines and seven fetal tissues (see supplementary
Fig. S1). In two hybridization experiments weak but
reproducible STK33-specific signals were produced in a
small number of tissues. Significant hybridization sig-
nals were obtained in cDNA from testis, fetal lung,
fetal heart, pituitary gland and kidney. Weak but still

in single alveolar cells (Fig. 2D,F), which, as revealed
by nuclear staining, probably are alveolar macro-
phages. In situ RNA hybridization in mouse retina
showed a signal in the outer plexiform layer (Fig. 2G).
Protein distribution
To investigate protein distribution, a polyclonal anti-
body against a Stk33-specific synthetic peptide was
generated. The specificity of the anti-Stk33 antibody
was determined by competition tests with synthetic
Stk33-specific peptide, anti-Stk33 signal disappeared in
testis sections, when antibody is preabsorbed with
12.5 ngÆlL
)1
synthetic peptide (Fig. 3, C3). Immuno-
staining on tissue sections with the Stk33 antibody
(Fig. 4) was observed in the same regions as the
mRNA in situ-hybridization (Fig. 2). In testis an
intensive staining was observed in only few cells per
tubulus, which may be classified as secondary sperma-
tocytes according to nucleus morphology and localiza-
tion (Figs 4 and 5). Round spermatides showed signals
of lower intensity compared to the spermatocytes.
Moreover, Stk33 positive and negative spermatides
were often seen in groups restricted to distinct tubular
profiles (Fig. 4A). Immunostaining signal was also vis-
ible in Sertoli cells, concentrated in the perinuclear
space. In all cases, the protein localization appeared to
be cytoplasmic.
In lung, immunostaining with anti-Stk33 antibody
produced strong signals in epithelial cells and suppo-

(dpp) mice, whereas 10 dpp were Stk33 negative (see
Fig. 3D). Although the Western blot results were not
A. O. Mujica et al. STK33 expression pattern
FEBS Journal 272 (2005) 4884–4898 ª 2005 FEBS 4887
evaluated quantitatively, it seems obvious that the sig-
nal is stronger in older mice.
Discussion
Here we report the first survey of the distribution of the
recently discovered serine ⁄ threonine kinase 33 in mouse
and human tissues. Our results are well in accordance
with the STK33 ⁄ Stk33 expression pattern derived from
the expressed sequence databases (compiled in Table 1).
The predominant expression of STK33 ⁄ Stk33 in testis
is also observable in UniGene EST database [32]
(http://www.ncbi.nlm.nih.gov/UniGene), Gene Expres-
sion Atlas [33] (http://symatlas.gnf.org/SymAtlas/) and
Testis
A
B C
Lung
Retina
D
G
E
F
Fig. 2. Distribution of Stk33 mRNA in testis, lung and retina demonstrated by in situ hybridization (ISH). (A,B) ISH in testis. Strong Stk33 signal
is detected in spermatocytes (spc), spermatides (sd) and possibly in spermatogonia (sg). No signal is detected around Sertoli cell nucleus (sen)
or in spermatozoa (sz), neither in Leydig cells nor any kind of cell in the interstitial space (is). (C) Nuclear staining with DAPI was used for char-
acterization of the nuclei in all tissues and is shown here exemplary for testis. (D,E,F) ISH in lung. Stk33 signal was detected in epithelium (ep)
and alveolar macrophages (am). No signal was found in cartilage (ca), smooth muscle (sm), connective tissue (ct), bronchioli (bri), aleveolar

by ISH and immunostaining. Additionally, Stk33
RNA ⁄ protein is expressed in some regions of the ner-
vous system in fetal mice (Fig. 6), and protein distribu-
tion in the adult brain shows a distinct distribution
that will be presented in a separate paper.
Human fetal lung yielded the second highest signal in
our cDNA dot-blot hybridization, whereas there was
remarkably weaker or no signal in human adult lung
samples. Northern blot analysis with mRNA also
showed no signal in adult mice lung, even though more
mRNA was immobilized from lung than from testis.
On the other hand, both mRNA in situ hybridization
and immunostaining experiments on tissue slices of
mouse adult lung showed a reproducible signal in bron-
chial epithelium and putative alveolar macrophages. It
seems evident that low expression of Stk33 is difficult
to examine by hybridization methods such as northern
blot and cDNA dot-blots using whole organs in all
tissues but testis. We were not able to detect Stk33
protein in mouse fetal lung. As we tested embryos
only at 15 days postcoitus, it is conceivable that expres-
sion in fetal lung occurs at different developmental
stages.
According to their predicted general biochemical
features, STK33 and Stk33 are probably soluble pro-
teins. psort analysis [36] found no notable known sig-
nal in STK33 ⁄ Stk33 primary structure except from
conserved C-terminal di-lysine motifs (511-Thr-Lys-
Lys-Lys-514 in human and 488-Gly-Lys-Lys-Arg-491
in mouse), which are recognized as putative endoplas-

(C2) and 12.5 ngÆlL
)1
(C3). C2 and C3
were photographed with longer exposure times and accordingly
exhibit the light-coloured regions in the interstitial space also seen
in negative control (C4) with no primary antibody. (D) Stk33 sper-
matogenesis specific expression demonstrated by anti-Stk33 anti-
body staining. Immunoreactivity is observable in recombinant
protein (Rec), protein extracts from testis of mice at 20 and 30 dpp
but absent in 10 dpp. Coomassie blue staining of total protein
shows the homogeneous presence of 75 lg protein extract in each
track. Recombinant protein is not visible in the Coomassie blue
stained control track (Rec), as the amount loaded (80 ng) is below
the detection threshold [48].
A. O. Mujica et al. STK33 expression pattern
FEBS Journal 272 (2005) 4884–4898 ª 2005 FEBS 4889
heart (Fig. 6B) and the lack of signal in adult mouse
heart in our northern blot analysis (Fig. 1A). A study
on human heart malformations using DNA micro-
arrays [38] revealed STK33 among the most downregu-
lated genes in patients with tetralogy of Fallot, a
nonfatal congenital cardiovascular malformation in
which ventricles are not fully separated by the septum
and the pulmonary artery valve is narrowed, causing a
partial mixing of venal and arterial blood and a
decrease of blood flow to the lungs. On the other hand,
data from the source database [34] suggests a higher
expression of STK33 in neuroblastoma than in any
Testis
AB C

expression pattern may reflect a cell division stage-spe-
cific synchrony of Stk33 expression during spermato-
genesis. This observation is strongly supported by the
Western blot results showing no signal in 10 dpp mice
and positive signal first by 20 dpp. The absence of sig-
nal at 10 dpp excludes involvement of Stk33 in Sertoli
cell or spermatogonia proliferation, which extends to
12 dpp [41]; but the signals at 20 and 30 dpp suggest a
meiosis and ⁄ or spermiogenesis specific function which
first occur at the periods of 8–18 dpp and 18–30 dpp
[41]. It has been hypothesized that novel cell division
regulatory checkpoints probably exist in the germ line
of higher eukaryotic organisms which are not necessar-
ily present in the basic ones already described for yeast
[42]. On the other hand, STK33 orthologs are present
in the genomes of several chordate organisms (Fig. 7),
but not found in yeast, fly or nematode genomes. This
is not too surprising as humans possess 74 members of
the CAMK group of protein kinases whereas only 21
are detectable in yeast, 32 in fly and 46 in worm [8].
We propose that STK33 ⁄ Stk33 expression occurs only
within a narrow time window during the spermatogen-
esis. It remains to be established whether this ‘pulse
like’ expression is directly associated with some germ
cell development checkpoint, which based on our data
seems suggestive, or rather reflects synchrony to other
events of the spermatogenesis. For instance, the mech-
anisms unleashing the pass of the haploid germ cells
through the tight junctions from the adluminal to the
luminal region are still unknown. It has been argued

A
B
C
D
E
G
H
Fig. 5. Stk33 in mouse secondary spermatocytes. Double staining
with anti-Stk33 IgG (red) and nuclear staining with DAPI (green). (A)
Detail of a section of two seminiferous tubules showing a single
Stk33-positive cell. Note the dashed line marking the borders
between two different tubuli sections. The DAPI staining reveals
the different nuclear morphologies of: sertoli cell nucleous (sen),
spermatogonia (sg), primary spermatocytes (sc1), secondary sper-
matocytes (sc2), round spermatides (rsd) and spermatozoa (sz).
According to their chromatine condensation, Stk33-positive sperma-
tocytes were found in metaphase II (A), prophase II (B,C,D) and
anaphase II (E,F,G, chromosome segregation indicated by arrows).
A. O. Mujica et al. STK33 expression pattern
FEBS Journal 272 (2005) 4884–4898 ª 2005 FEBS 4891
and 42 °C +50% v ⁄ v formamide for northern blots).
After several washes with 2 · NaCl ⁄ Cit and 0.1 NaCl ⁄
Cit at room temperature membranes were exposed to
X-ray film with or without intensifying screens at
)70 °C. Exposure time varied from several hours to sev-
eral days.
Fig. 6. Localization of Stk33 in mouse embryo, day 15 postcoitius. (A) Overview of the mouse embryo immunostaining analysed with non-
confocal laser scanning. Regions with particular strong signal (a1–a3 and b) were photographed with the fluorescence microscope. Signal is
detected in some regions in the head, for instance between pons and medulla oblongata (A1), intermediate zone of mesencephalon (A2)
and medulla (A3). Strong signal is observed in heart ventricle (B) in particular in endocardium (B1–B3), here supported by nuclear staining in

[45]. mRNA was isolated from total RNA with the Nucleo-
Trap
Ò
isolation kit (Macherey-Nagel, Du
¨
ren, Germany),
following the manufacturer’s instructions. mRNA was
quantified spectrophotometrically, and separated by electro-
phoresis on a 1.2% agarose gel under denaturing conditions
(5.5% formaldehyde), transferred to Nylon membranes
(Roche, Mannheim, Germany) and UV-cross linked. The
blot was hybridized with a DNA Stk33-specific probe and
a probe from the L19 housekeeping gene as normalization
control. BioMax MS autoradiography films were exposed
to the radioactive blot with intensifying screen at )70 °C.
Exposure time was determined empirically.
Fig. 7. Amino acid sequence alignment of
Stk33 kinase domain in some chordates.
Proteins sequences from human (Hsa) and
mouse (Mmu) as described in [1].
Sequences from Rattus norvegicus (Rno),
Fugu rubripes (Fru), Danio rerio (Dre) and
Ciona intestinales (Cin), were inferred from
their respective genome projects already
available [49–51]. Partial Stk33 EST
sequences are detected in Xenopus laevis
and X. tropicalis, but they still do not extend
the whole kinase domain and were in con-
sequence excluded from the analysis. The
putative ATP-binding subdomain, serine ⁄

paraformaldehyde solution [46]. The right atrium was
opened to enable venous outflow. Organs were prepared,
cryoprotected in a gradient of sucrose and stored in
NaCl ⁄ P
i
)30% sucrose. Cryosections (8–15 lm) were pre-
pared and mounted onto glass slides coated with silane.
Additionally, some tissues and 15-day embryos of sacrificed
Balb ⁄ C mice were taken, fresh frozen on dry ice and stored
at )80 °C. Cryosections of these tissues (8–15 lm) were
fixed in 4% paraformaldehyde–NaCl ⁄ P
i
for 10 min prior to
immunohistochemistry.
RNA in situ hybridization
Templates for the production of sense and antisense RNAs
were produced by adding T7 viral RNA polymerase pro-
moter adaptors to Stk33-PCR amplification products fol-
lowing standard procedures [47]. Accordingly, template for
the production of sense RNA was amplified with the
primers 5¢-CAGAGATGCATAATACGACTCACTATAGG
GAGAAACCCAGAAAGTGATGAG-3¢ and 5¢-TAGAA
CTAAGCGAGCATG-3¢, whereas the template for the
production of the antisense strand was amplified using the
primers 5¢-CAGAGATGCATAATACGACTCACTATAG
GGAGATAGAACTAAGCGAGCATG-3¢ and 5¢-AACC
CAGAAAGTGATGAG-3¢. The probes were used as tem-
plates for in vitro RNA synthesis and labelling with
digoxigenin (RNA in vitro transcription from Roche) fol-
lowing the manufacturer’s instructions. Briefly, RNA

services/TMHMM/) and tmpred (http://www.ch.embnet.
org/software/TMPRED_form.html). It is likely to be exter-
nally oriented according to sspro2 (http://promoter.ics.
uci.edu/BRNN-PRED) and blastp searches with low strin-
gency parameters (expected value ¼ 1000, word size ¼ 2)
against human and mouse protein databases underline its
high Stk33 specificity (http://www.ncbi.nlm.nih.gov/blast).
The peptide was synthesized adding a cysteine residue in
the N terminal for keyhole limpet haemocyanin conjuga-
tion. Two rabbits were immunized with the keyhole limpet
haemocyanin-conjugated peptide for 10 weeks. Peptide syn-
thesis and rabbit immunization were performed by Gene-
med Synthesis (San Francisco, CA, USA).
The anti-Stk33 polyclonal serum was purified by affinity
chromatography using immobilized sulfhydryl-containing
Stk33-peptide to SulfoLink coupling gel (Pierce Biotechno-
logy, Rockford, IL, USA). Coupling, blocking, prepar-
ation of the column and purification of the antiserum
were carried out following manufacturer’s instructions.
Antibody was eluted from the column with glycine buffer
(100 mm, pH 2.5). Fractions were neutralized by adding
1 m Tris pH 9. The affinity-purified antibody is in a final
concentration of  25 lgÆmL
)1
and preserved in 0.05%
sodium azide.
The activity of the antibody was tested by western blot-
ting with a recombinant biotin-Stk33 fusion protein
expressed in E. coli (Fig. 3B). In our hands, anti-Stk33
diluted 1 : 40 (0.625 lgÆ mL

paste, followed by sonication on ice (Branson Sonifier Cell
Disruptor B15). The lysate was centrifugated at 10 000 g
for 15 min at 4 °C to remove cellular debris. The biotinyl-
ated Stk33-fusion protein was affinity-purified using Soft
Link
TM
Soft Release Avidin Resin (Promega) following the
manufacturer’s instructions.
Western blotting
Recombinant Stk33-Protein was separated on 12%
SDS ⁄ polyacrylamide gels and electrophoretically trans-
ferred to nitrocellulose membranes (Schleicher & Schuell,
Dassel, Germany) using 48 mm Tris, 39 mm glycine,
0.037% SDS pH 8.3 containing 20% methanol. Following
transfer, the nitrocellulose membrane was incubated in
blocking buffer (NaCl ⁄ P
i
containing 3% BSA fraction V,
Sigma, St Louis, MO, USA) for at least 1 h at 4 °C,
washed in TBS and then incubated with the anti-Stk33 anti-
body for at least 2 h at room temperature with gentle agita-
tion. The membrane was rinsed with TBS (five times for
5 min each), incubated with alkaline phosphatase-conju-
gated AffiniPure goat Anti-Rabbit IgG (H + L) (Dianova,
Hamburg, Germany) for 1 h at room temperature and
washed three times for 5 min with TBS and twice for 5 min
with buffer 100 mm Tris, 100 mm NaCl, 50 mm MgCl
2
pH 9.5. Bound anti-Stk33 antibodies were detected by
staining with NBT ⁄ BCIP solution (Roche).

3
cell, BIO RAD Laboratories). Separated proteins were
electrophoretically transferred (Mini Trans-Blot
Ò
Electro-
phoretic Transfer Cell, BIO RAD Laboratories) to nitro-
cellulose membranes (Schleicher & Schuell) using 48 mm
Tris, 39 mm glycine, 0.037% SDS, pH 8.3 containing 20%
methanol. Following transfer, the nitrocellulose membrane
was incubated in blocking buffer (NaCl/P
i
-T: phosphate-
buffered saline (NaCl/P
i
), 0.1% Tween-20 containing 5%
nonfat dry milk [Applichem, Darmstadt, Germany]) for at
least 1 h at room temperature, washed twice briefly in
NaCl ⁄ P
i
-T and then incubated with the anti-Stk33 antibody
(1 : 100 dilution in NaCl ⁄ P
i
-T) overnight at room tempera-
ture with gentle agitation. The membrane was rinsed with
NaCl ⁄ P
i
-T (twice briefly, once for 15 min, three times for
5 min), incubated with monoclonal anti-rabbit IgG Peroxi-
dase conjugate (Sigma) for 2 h at room temperature and
washed with NaCl ⁄ P

fragment donkey anti-
rabbit IgG (Dianova) at 1 : 100 dilution in 10%
FCS ⁄ NaCl ⁄ P
i
for 1 h at room temperature. Double-stain-
ing experiments with the monoclonal anti-a-tubulin anti-
body (Sigma) sections were additionally fixed with 20%
methanol at )20 °C for 10 min and incubated at 1 : 7000
dilution in 10% FCS ⁄ NaCl ⁄ P
i
overnight at 4 °C. Detection
of a-tubulin was performed using fluorescein goat anti-
mouse IgG (Oncogene Research Products, San Diego, CA,
USA). Nuclei were stained with DAPI prior to washing
with NaCl ⁄ P
i
. Sections were mounted with 1 gÆL
)1
pheny-
lenediamine in 50% v ⁄ v phosphate-buffered glycerol.
Control incubations were carried out essentially as the
immunostaining reactions but omitting primary antibodies.
The specificity of the immunostaining was demonstrated
performing preabsorption tests of the Stk33-antibody with
synthetic peptide. Before applying it to the testis cryostat
sections, the antibody was diluted 1 : 40 in NaCl ⁄ P
i
, 10%
FCS, incubated overnight with different concentrations of
antigen (1.25 ng ÆlL

the mouse embryo slides, Christof Rickert for assist-
ance with the LSM and Oliver Bitz for assistance with
the Laser Scanner. Financial assistance was obtained
from the Rheinland-Pfalz Stiftung fu
¨
r Innovation and
through funding by the German Human Genome Pro-
ject (DHGP grant 01KW9624) which is gratefully
acknowledged.
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The following material is available for this article
online:
Fig. S1. Distribution of cDNAs in Clontech’s MTE
array, catalog # 7775–1, Protocol # PT3307-1, Version
# PR89545.
Fig. S2. Exemplary negative controls of RNA Stk33 in
situ hybridization and immunostaining in testis.
Fig. S3. Immunohistochemistry with anti-Stk33 and
negative controls without primary antibody of mouse
embryos.
STK33 expression pattern A. O. Mujica et al.
4898 FEBS Journal 272 (2005) 4884–4898 ª 2005 FEBS