Won Heo

Identification of a Novel Divergent Calmodulin Isoform from Soybean Which Has Differential Ability to Activate Calmodulin-dependent Enzymes

Calmodulin plays pivotal roles in the transduction of various Ca-mediated signals and is one of the most highly conserved proteins in eukaryotic cells. In plants, multiple calmodulin isoforms with minor amino acid sequence differences were identified but their functional significances are unknown. To investigate the biological function of calmodulins in the regulation of calmodulin-dependent enzymes, we cloned cDNAs encoding calmodulins in soybean. Among the five cDNAs isolated from soybean, designated as SCaM-1 to −5, SCaM-4 and −5 encoded very divergent calmodulin isoforms which have 32 amino acid substitutions from the highly conserved calmodulin, SCaM-1 encoded by SCaM-1 and SCaM-3. SCaM-4 protein produced in Escherichia coli showed typical characteristics of calmodulin such as Ca-dependent electrophoretic mobility shift and the ability to activate phosphodiesterase. However, the extent of mobility shift and antigenicity of SCaM-4 were different from those of SCaM-1. Moreover, SCaM-4 did not activate NAD kinase at all in contrast to SCaM-1. Also there were differences in the expression pattern of SCaM-1 and SCaM-4. Expression levels of SCaM-4 were approximately 5-fold lower than those of SCaM-1 in apical and elongating regions of hypocotyls. In addition, SCaM-4 transcripts were barely detectable in root whereas SCaM-1 transcripts were as abundant as in apical and elongating regions of hypocotyls. In conclusion, the different biochemical properties together with differential expression of SCaM-4 suggest that this novel calmodulin may have different functions in plant cells.

Identification of a Calmodulin-Regulated Soybean Ca2+-ATPase (SCA1) That Is Located in the Plasma Membrane

Ca2+-ATPases are key regulators of Ca2+ ion efflux in all eukaryotes. Animal cells have two distinct families of Ca2+ pumps, with calmodulin-stimulated pumps (type IIB pumps) found exclusively at the plasma membrane. In plants, no equivalent type IIB pump located at the plasma membrane has been identified at the molecular level, although related isoforms have been identified in non–plasma membrane locations. Here, we identify a plant cDNA, designated SCA1 (for soybean Ca2+-ATPase 1), that encodes Ca2+-ATPase and is located at the plasma membrane. The plasma membrane localization was determined by sucrose gradient and aqueous two-phase membrane fractionations and was confirmed by the localization of SCA1p tagged with a green fluorescent protein. The Ca2+-ATPase activity of the SCA1p was increased approximately sixfold by calmodulin (K1/2 ∼10 nM). Two calmodulin binding sequences were identified in the N-terminal domain. An N-terminal truncation mutant that deletes sequence through the two calmodulin binding sites was able to complement a yeast mutant (K616) that was deficient in two endogenous Ca2+ pumps. Our results indicate that SCA1p is structurally distinct from the plasma membrane–localized Ca2+ pump in animal cells, belonging instead to a novel family of plant type IIB pumps found in multiple subcellular locations. In plant cells from soybean, expression of this plasma membrane pump was highly and rapidly induced by salt (NaCl) stress and a fungal elicitor but not by osmotic stress.

Competitive binding of calmodulin isoforms to calmodulin-binding proteins: implication for the function of calmodulin isoforms in plants

In plants, multiple calmodulin (CaM) isoforms exist in an organism which vary in their primary structures in as much as 32 residues out of their 148 amino acids. These CaM isoforms show differences in their expression patterns and/or target enzyme activation ability. To further understand the biological significance of CaM isoforms, we examined whether CaM isoforms act on specific regulatory targets. In gel overlay assays on various soybean tissue extracts, surprisingly, two soybean CaM isoforms (SCaM-1 and SCaM-4) did not show significant differences in their target binding protein profiles, although they exhibited minor differences in their relative target binding affinities. In addition, both SCaM isoforms not only effectively bound five known plant CaMBPs, but also showed competitive binding to these proteins. Finally, immunolocalization experiments with the SCaM proteins in sections of various tissues using specific antibodies revealed similar distribution patterns for the SCaM isoforms except for root tissues, which indicates that the SCaM isoforms are concomitantly expressed in most plant tissues. These results suggest that CaM isoforms may compete for binding to CaMBPs in vivo. This competitive nature of CaM isoforms may allow modulation of Ca(2+)/CaM signaling pathways by virtue of relative abundance and differential target activation potency.

Is Titanium Mesh Cage Safe in Surgical Management of Pyogenic Spondylitis

OBJECTIVEnTo report our experience with pyogenic spondylitis treated with anterior radical debridement and insertion of a titanium mesh cage and to demonstrate the effectiveness and safety of the use of a titanium mesh cage in the surgical management of pyogenic spondylitis.nnnMETHODSnWe retrospectively analyzed the clinical characteristics of 19 patients who underwent surgical treatment in our department between January 2004 and December 2008. The average follow-up period was 11.16 months (range, 6-64 months). We evaluated risk factors, cultured organisms, lab data, clinical outcomes, and radiographic results. Surgical techniques for patients with pyogenic spondylitis were anterior radical debridement and reconstruction with titanium mesh cage insertion and screw fixation. All patients received intravenous antibiotics for at least 6 weeks postoperatively, and some patients received oral antibiotics.nnnRESULTSnThe infections resolved in all of the patients as noted by normalization of their erythrocyte sedimentation rates and C-reactive protein levels. The mean pain score on a Visual Analog Scale was 7.8 (range, 4-10) before surgery and 2.4 (range, 1-5) after surgery. The Frankel grade was improved by one grade in seven patients. After surgery, the average difference of the angle was improved about 6.96° in all patients. At the last follow-up, the mean loss of correction was 4.86°.nnnCONCLUSIONnAnterior radical debridement followed by the placement of instrumentation with a titanium mesh cage may be a safe and effective treatment for selected patients with pyogenic spondylitis. This surgical therapy does not lead to recurrent pyogenic spondylitis.

Cystic Meningiomas: Correlation between Radiologic and Histopathologic Features

Background Tumors with cysts often correlate with gliomas, metastatic tumors, or hemangioblastomas, which require differentiation. Methods Thirty-eight cases of cyst associated-meningioma based on preoperative radiologic studies and histologic confirmations were reviewed from November 1998 to July 2017. Results A total of 395 cases of meningioma were observed in the 20 years, and surgical treatment of intracranial meningioma was performed in 120 cases. Thirty-eight (9.6%) cases of cyst associated meningiomas were analyzed. Nauta type I was the most common type of cyst (39.5%) and the most frequent histopathological subtype was meningothelial type (36.8%). Conclusion Statistically there were no significant associations between meningioma histopathological type and associated cysts; however, the rate of World Health Organization grade II was higher in cyst associated meningiomas than in unrelated meningiomas. This correlation was weak, in accordance with the meningioma grade.

Lateral Lumbar Interbody Fusion and in Situ Screw Fixation for Rostral Adjacent Segment Stenosis of the Lumbar Spine

Objective The purpose of this study is to describe the detailed surgical technique and short-term clinical and radiological outcomes of lateral lumbar interbody fusion (LLIF) and in situ lateral screw fixation using a conventional minimally invasive screw fixation system (MISF) for revision surgery to treat rostral lumbar adjacent segment disease. Methods The medical and radiological records were retrospectively reviewed. The surgery was indicated in 10 consecutive patients with rostral adjacent segment stenosis and instability. After the insertion of the interbody cage, lateral screws were inserted into the cranial and caudal vertebra using the MISF through the same LLIF trajectory. The radiological and clinical outcomes were assessed preoperatively and at 1, 3, 6, and 12 months postoperatively. Results The median follow-up period was 13 months (range, 3–48 months). Transient sensory changes in the left anterior thigh occurred in 3 patients, and 1 patient experienced subjective weakness; however, these symptoms normalized within 1 week. Back and leg pain were significantly improved (p<0.05). In the radiological analysis, both the segmental angle at the operated segment and anterior disc height were significantly increased. At 6 months postoperatively, solid bony fusion was confirmed in 7 patients. Subsidence and mechanical failure did not occur in any patients. Conclusion This study demonstrates that LLIF and in situ lateral screw fixation may be an alternative surgical option for rostral lumbar adjacent segment disease.