What is new in the treatment of hypospadias?

Hypospadias is a congenital anomaly characterized by a ventrally placed urethral meatus in a more proximal position on the midline than its normal position in the glanular part of the penis. In 1961, C. E. Horton and C. J. Devine, Jr., developed single-stage modern surgical techniques, namely, local skin flaps and free skin grafts, for urethra reconstruction in hypospadias repair, which may be applied to almost any case with different localizations of the meatus. Later, two new methods, advancement of the urethra and preputial island flap techniques, were added to the surgical algorithm. Because acceptable results were observed, the authors have insisted on using these four techniques for all hypospadias cases since 1972. Complication rates (mainly fistula formation) were quite high (50 percent) in their early series of adults as a result of erection and hematoma formation. The complication rate of their patient population, which is now mainly composed of preschool children, has decreased to 7 to 8 percent, primarily as a result of careful selection of appropriate techniques for each individual case, the development of better surgical materials and equipment, and taking necessary precautions for postoperative care. A brief summary of modern hypospadias repair techniques is presented in four major classes. The results of the authors' 30-year experience and the precautions necessary to avoid postoperative complications are evaluated. The authors conclude that the four modern techniques and their modifications should be performed meticulously for successful hypospadias repair.     

What is Causal Specificity About,and What is it Good for in Philosophy of Biology?

Acta Biotheoretica - The concept of causal specificity is drawing considerable attention from philosophers of biology. It became the rationale for rejecting (and occasionally, accepting) a thesis...     

What is signal and what is noise in the brain?

The response of a cortical neuron to a stimulus can show a very large variability when repeatedly stimulated by exactly the same stimulus. This has been quantified in terms of inter-spike-interval (ISI) statistics by several researchers (e.g., [Softky, W., Koch, C., 1993. The highly irregular firing of cortical cells is inconsistent with temporal integration of random EPSPs. J. Neurosci. 13(1), 334-350.]). The common view is that this variability reflects noisy information processing based on redundant representation in large neuron populations. This view has been challenged by the idea that the apparent noise inherent in brain activity that is not strictly related or temporally coupled to the experiment could be functionally significant. In this work we examine the ISI statistics and discuss these views in a recently published model of interacting cortical areas [Knoblauch, A., Palm, G., 2002. Scene segmentation by spike synchronization in reciprocally connected visual areas. I. Local effects of cortical feedback. Biol. Cybernet. 87(3), 151-167.]. From the results of further single neuron simulations we can isolate temporally modulated synaptic input as a main contributor for high ISI variability in our model and possibly in real neurons. In contrast to alternative mechanisms, our model suggests a function of the temporal modulations for short-term binding and segmentation of figures from background. Moreover, we show that temporally modulated inputs lead to ISI statistics which fit better to the neurophysiological data than alternative mechanisms.     

Is membrane occupation and recognition nexus domain functional in plant phosphatidylinositol phosphate kinases?

Phosphatidylinositol phosphate kinase (PIPK) catalyzes a key step controlling cellular contents of phosphatidylinositol-4,5-bisphosphate [PtdIns(4,5)P₂], a critical intracellular messenger involved in vesicle trafficking and modulation of actin cytoskeleton and also a substrate of phospholipase C to produce the two intracellular messengers, diacylglycerol and inositol-1,4,5-trisphosphate. In addition to the conserved C-terminal PIPK catalytic domain, plant PIPKs contain a unique structural feature consisting of a repeat of membrane occupation and recognition nexus (MORN) motifs, called the MORN domain, in the N-terminal half. The MORN domain has previously been proposed to regulate plasma membrane localization and phosphatidic acid (PA)-inducible activation. Recently, the importance of the catalytic domain, but not the MORN domain, in these aspects was demonstrated. These conflicting data raise the question about the function of the MORN domain in plant PIPKs. We therefore performed analyses of PpPIPK1 from the moss Physcomitrella patens to elucidate the importance of the MORN domain in the control of enzymatic activity; however, we found no effect on either enzymatic activity or activation by PA. Taken together with our previous findings of lack of function in plasma membrane localization, there is no positive evidence indicating roles of the MORN domain in enzymatic and functional regulations of PpPIPK1. Therefore, further biochemical and reverse genetic analyses are necessary to understand the biological significance of the MORN domain in plant PIPKs.Key words: membrane occupation and recognition nexus (MORN) domain, phosphatidylinositol phosphate kinase, phosphatidic acid, Physcomitrella patensPhosphoinositides (PIs) are minor membrane phospholipds that play pivotal roles in various signal transduction cascades involved in development and stress response via the regulation of cytoskeletal organization, ion channel activation and vesicle trafficking.^1,2 These are derivatives of phosphatidylinositol (PtdIns) produced by phosphorylation of the 3-, 4- and 5- positions of the inositol ring.² To address the roles of PIs, enzymes involved in their production have been extensively studied using biochemical and molecular biological approaches. Of these enzymes, phosphatidylinositol monophosphate kinases (PIPKs) catalyze the reaction producing phosphatidylinositol-4,5-bisphosphate [PtdIns(4,5)P₂] that is a substrate of phospholipase C and phosphatidylinositol 3-kinase, and also acts as an intracellular messenger involved in the regulation of F-actin organization and activity of ion channels.^1–3 Although PtdIns(4,5)P₂ is produced by sequential phosphorylation by phosphatidylinositol 4-kinase, producing phosphatidylinositol-4-phosphate [PtdIns(4)P], and then by PIPK,^1,2 the cellular levels of PtdIns(4)P are much higher compared to PtdIns(4,5)P₂.^4–6 Thus, a restriction step controlling cellular PtdIns(4,5)P₂ contents is mediated by PIPKs, indicating the importance of PIPK regulation in various kinds of physiological processes.The roles of plant PIPKs have been established in growth regulation, such as polarized tip growth of root hairs and pollen tubes, via their localization at plasma membranes.^7–12 It is worth to note that plant PIPKs contain a unique structure consisting of a repeat of a membrane occupation recognition nexus (MORN) motifs, called MORN domain, at the N-terminal region and a C-terminal PIPK catalytic domain, except for AtPIP5K10 and AtPIP5K11 from Arabidopsis thaliana, which lack the N-terminal MORN domain.¹³ The MORN domain was first identified as plasma membrane-binding module in junctophilin¹⁴ and the involvement of the MORN domain in plasma membrane localization was proposed for A. thaliana AtPIP5K1 and AtPIP5K3.^9,15,16Another remarkable feature of eukaryotic PIPKs is dependency of the enzymatic activity on phosphatidic acid (PA).^17,18 Indeed, PA-dependent activation of PIPKs has been observed in A. thaliana and in the moss Physcomitrella patens,^6,19,20 as with animal type I PIPKs.²¹ Although much less is known about how PA activates PIPKs in plants, biochemical analyses suggested the involvement of the MORN domain in PA-dependent activation of AtPIP5K1.¹⁵Based on above findings, it was proposed that plasma membrane-localization and PA-dependent activation of plant PIPKs might be regulated by the MORN domain.^9,15,16 In contrast, we recently demonstrated the critical involvement of the C-terminal half containing the catalytic domain of plant PIPKs in both plasma membrane-localization and PA-dependent activation.²² Thus, the function of the MORN domain remains elusive in plant PIPKs.As shown earlier, the N-terminal half of P. patens PpPIPK1 containing the MORN domain enhances its catalytic activity.²² Thus, to identify the region required for the activation of PpPIPK1, we further dissected the N-terminal half into 3 regions; the N-terminal region (amino acid nos. 1–154), the MORN repeat (amino acid nos. 155–316) and the linker region (amino acid nos. 338–452), and made deletion mutants of PpPIPK1 as shown in Figure 1A. Using Pfu Turbo DNA polymerase (Stratagene, La Jolla, USA), DNA fragments corresponding to deletion mutants lacking the N-terminal and N-terminal plus the MORN repeat, designated PpPIPK1ΔN and PpPIPK1ΔN-MORN, respectively, were amplified with primer sets; one is M_PIPK1_fb (5′-GGC AAG CAC GTG TAT AAT GTC TGA AGG GCT T-3′) and XhoIPIPK1 (5′-TAA ACT CGA GTT AGC TGG GTA GGA GGA AA-3′) and the other is M_PIPK1_f7 (5′-AGA GAA CAC GTG TAT AAT GTC TGA CTT CTA CGT CGG T-3′) and XhoIPIPK1. For building an expression plasmid for a deletion mutant lacking the MORN repeat, designated PpPIPK1ΔMORN, the N-terminal region and PpPIPK1ΔN-MORN were amplified with primer sets, M_PIPK1_fb and M_PIPK1_r3 (5′-TTG TAA GTC TCG GGT GCC ATT TGA GAG CTC-3′) M_PIPK1_f6 (5′-GAG CTC TCA AAT GGC ACC CGA GAC TTA CAA-3′) and XhoIPIPK1, respectively, using Pfu Turbo DNA polymerase and resultant DNA fragments were fused by PCR with a primer set, M_PIPK1_fb and XhoIPIPK1 using the same enzyme. These PCR products were digested with Pml1 and XhoI and inserted into Pml1-XhoI digested pPICZB (Invitrogen) to construct expression plasmids, pPICZB-PpPIPK1ΔN, pPICZB-PpPIPK1ΔN-MORN and pPICZB-PpPIPK1ΔMORN. Transformation of P. pastoris X-33 cells with the above expression plasmids, colony PCR of transformants and following expression, purification and western blot analysis of His-tagged recombinant proteins were performed as described previously.⁶ The PIPK activity assay using purified His-tagged proteins was carried out as described previously²³ with the modifications.⁶Open in a separate windowFigure 1Functional dissection of the N-terminal region of PpPIPK1 identifies positive regulatory regions. (A) His-tagged recombinant PpPIPK1 proteins. A repetition of eight MORN motifs (grey boxes) and the conserved catalytic domain (black box) are indicated in wild type and mutant PpPIPK1s. The MORN repeat and junction of internal deletion are indicated by amino acid position numbers. (B) In vitro lipid kinase activity of His-tagged recombinant proteins. The activities of recombinant proteins bound to Ni-NTA agarose beads were assayed with PtdIns4P. (C) In vitro PA-dependent lipid kinase activity of His-tagged proteins. The activities of recombinant proteins bound to Ni-NTA agarose beads were assayed with PtdIns4P with 143 µM PA. Top and bottom arrowheads represent reaction products PtdIns(4,5)P₂ and lysoPtdIns(4,5)P₂, respectively.Biochemical analyses of these enzymes after expression in yeast P. pastoris X-33 cells followed by purification showed that deletion of the N-terminal region (PpPIPK1ΔN) reduced PpPIPK1 activity ca 40% compared to the full length enzyme, whereas loss of the MORN repeat (PpPIPK1ΔMORN) had no significant effect (Fig. 1B). In agreement, a mutant lacking four MORN repeats of the total eight repeats showed no difference in the activity compared the full length enzyme (data not shown). These results indicate a positive role of the N-terminal region, but not the MORN repeats, on PpPIPK1 activity. However, these findings differ from those obtained with AtPIP5K1, where the MORN domain represses enzymatic activity.¹⁵ Interestingly, PpPIPK1ΔN-MORN containing the linker and catalytic regions showed higher enzymatic activity of ca 23 % compared to the full length PpPIPK1 (Fig. 1B). The C-terminal half only containing the catalytic domain of PpPIPK1 and thus lacking the linker region showed a reduced activity.²² It is therefore proposed that the linker region carries a positive regulatory element. Although details are unknown, negligible effects of the N-terminal and MORN domains for the enzymatic activity has been indicated in AtPIP5K3 from A. thaliana.¹¹ Moreover, it is noteworthy that PA-dependent activation was not affected by any deletion as shown in Figure 1C, confirming that the N-terminal half is not involved in PA dependency of the PpPIPK1 activity.²²Our results indicated that the MORN domain is not involved in the regulation of the catalytic activity in PpPIPK1. Similarly, the function of the MORN domain found in the accumulation and replication of chloroplasts 3 (ARC3) was not resolved. ARC3 is an FtsZ homologue involved in chloroplast division²⁴ and the only protein containing the MORN repeats other than PIPKs in A. thaliana. It was shown that the ARC3 MORN domain did not interact with any stromal plastid division components.²⁵ Moreover, there are reports representing functions of the MORN domain other than plasma membrane binding. Human amyotrophic lateral sclerosis 2 (ALS2), a guanine nucleotide exchange factor (GEF) specific to the small GTPase Rab5, contains the MORN domain at the central region that is essential for the GEF activity but not for interaction with Rab5.²⁶ In contrast, specific interaction of the MORN domain with Rab-E GTPases and resultant enzymatic activation was recently demonstrated for AtPIP5K2.¹² It is interesting that these results are inconsistent with each other in terms of interaction of the MORN domain with small GTPases.Taken together, with no function of the MORN domain in plasma membrane localization of PpPIPK1 and AtPIP5K1,²² the function of the MORN domain is still unknown, despite its high conservation plants PIPKs. Alternatively, based on the findings of ARC3, ALS2 and AtPIP5K2,^12,25,26 the function of the MORN domain possibly varies among PIPK isoforms and may thus have multifunctional roles. Therefore, it is necessary to identify interaction partners for the MORN domain of each plant PIPKs and to analyze phenotypes of transgenic plants carrying MORN domain-lacking PIPKs during developmental process and environmental stress responses.     

What is in a model?

After reading contradictory claims of model status for some insect species, we feel a brief discussion of the topic may be useful. Here, we document a few examples where clarity on model status seems to be lacking, briefly review work on widely recognized models, and offer criteria for including any given species as a model organism.